Dr. Thavrak Huon from South Korea

[28] Detection, assay, and isolation of allene oxide

2010-02-25T13:25:00.003+09:00

The enzyme(s) now commonly referred to as allene oxide synthase(AOS), CYP74 was reported in 1966 as an activity in flaxseed that metabolizedlinoleic acid hydroperoxide to an alfa-ketol derivative. Early publicationsestablished that other associated products from unsaturated fattyacid hydroperoxides are y-ketols and cyclopentenone fatty acids. Thesederivatives are now recognized to arise by hydrolysis and rearrangementsof the initial enzymic product, the very unstable allene oxide. The early literature refers to AOS as "hydroperoxide isomerase '' or "hydroperoxide cyclase, ''and "hydroperoxide dehydrase" has also b e e nused...

Source: Methods in Enzymology (1996) Vol 272, p. s 250-259

CYP74C3 and CYP74A1, plant cytochrome P450 enzymes whose activity is regulated by ...

2009-12-21T20:54:00.005+09:00

CYP74C3 and CYP74A1, plant cytochrome P450 enzymes whose activity is regulated by detergent micelle association, and proposed new rules for the classification of CYP74 enzymes

CYP74C3 (cytochrome P450 subfamily 74C3), an HPL (hydroperoxide lyase) from Medicago truncatula (barrel medic), and CYP74A1, an AOS (allene oxide synthase) from Arabidopsis thaliana, are key membrane-associated P450 enzymes in plant oxylipin metabolism. Both recombinant detergent-free enzymes are monomeric proteins with dual specificity and very low enzyme activity that can be massively activated with detergent. This effect is a result of the formation of a complex between the protein monomer and a single detergent micelle and, in the case of CYP74A1, has a major effect on the substrate specificity of the enzyme. Association with a detergent micelle without an effect on protein oligomeric state represents a new mechanism of activation for membrane-associated P450 enzymes. This may represent a second unifying feature of CYP74 enzymes, in addition to their known differences in reaction mechanism, which separates them functionally from more classical P450 enzymes. Highly concentrated and monodispersed samples of detergent-free CYP74C3 and CYP74A1 proteins should be suitable for structural resolution. On the basis of recent evidence for incorrect assignment of CYP74 function, using the current rules for CYP74 classification based on sequence relatedness, we propose an alternative based on substrate and product specificity for debate and discussion.

Source: Biochem Soc Trans. (2006) vol. 34, p. 1223-1227

Preparative enzymatic solid phase synthesis of cis(+)-12-oxo-phytodienoic acid – physical interaction of AOS and AOC is not necessary

2009-11-17T20:41:00.005+09:00

The pathway of jasmonic acid (JA) biosynthesis was established in the 1980s by Vick and Zimmerman but, until now, the preparative biosynthesis of the jasmonic acid precursors 12-oxo-phytodienoic acid (OPDA) and 3-oxo-2-[2′-pentenyl]-cyclopentan-1-octanoic acid (OPC-8:0) in their endogenous and biologically relevant cis(+)-configuration was only possible in small amounts and had to put up with high costs. This was mainly due to the lack of high amounts of pure and enzymatically active allene oxide cyclase (AOC), which is a key enzyme in the biosynthesis of jasmonates in that it releases, in a coupled reaction with allene oxide synthase (AOS), the first cyclic and biological active metabolite – OPDA. We describe here the expression and purification of AOS and AOC and their subsequent coupling to solid matrices to produce an enantioselective, reusable bioreactor for octadecanoid production. With the method described here it is possible to produce optically pure enantiomers of octadecanoids in high amounts in a cost- and time-efficient manner. Furthermore, it could be demonstrated that a physical interaction of AOS and AOC, hitherto postulated to be required for substrate channeling from AOS to AOC, is not necessary for the in vitro cyclization of the unstable epoxide generated by the AOS reaction.

Keywords: Allene oxide cyclase; Allene oxide synthase; Jasmonates; Jasmonic acid; 12-Oxo-phytodienoic acid; Octadecanoid pathway; Oxylipins; Plant hormones

Source: Phytochemistry (2007) vol. 68, Issue 2, p. 229-236

Evidence for an Ionic Intermediate in the Transformation of Fatty Acid ...

2009-10-26T14:30:00.013+09:00

Evidence for an Ionic Intermediate in the Transformation of Fatty Acid Hydroperoxide by a Catalase-related Allene Oxide Synthase from the Cyanobacterium Acaryochloris marina

Allene oxides are reactive epoxides biosynthesized from fatty acid hydroperoxides by specialized cytochrome P450s or by catalase-related hemoproteins. Here we cloned, expressed, and characterized a gene encoding a lipoxygenase-catalase/peroxidase fusion protein from Acaryochloris marina. We identified novel allene oxide synthase (AOS) activity and a by-product that provides evidence of the reaction mechanism. The fatty acids 18.4ω3 and 18.3ω3 are oxygenated to the 12R-hydroperoxide by the lipoxygenase domain and converted to the corresponding 12R,13-epoxy allene oxide by the catalase-related domain. Linoleic acid is oxygenated to its 9R-hydroperoxide and then, surprisingly, converted ~70% to an epoxyalcohol identified spectroscopically and by chemical synthesis as 9R,10S-epoxy-13S-hydroxyoctadeca-11E-enoic acid and only ~30% to the 9R,10-epoxy allene oxide. Experiments using oxygen-18-labeled 9R-hydroperoxide substrate and enzyme incubations conducted in H218O indicated that ~72% of the oxygen in the epoxyalcohol 13S-hydroxyl arises from water, a finding that points to an ionic intermediate (epoxy allylic carbocation) during catalysis. AOS and epoxyalcohol synthase activities are mechanistically related, with a reacting intermediate undergoing a net hydrogen abstraction or hydroxylation, respectively. The existence of epoxy allylic carbocations in fatty acid transformations is widely implicated although for AOS reactions, without direct experimental support. Our findings place together in strong association the reactions of allene oxide synthesis and an ionic reaction intermediate in the AOS-catalyzed transformation.

Source: J. Biol. Chem. (2009) vol. 284, p. 22087-22098

Tomato CYP74C3 is a Multifunctional Enzyme not only Synthesizing Allene Oxide but also Catalyzing its Hydrolysis and Cyclization

2009-09-02T19:02:00.009+09:00

The mechanism of the recombinant tomato allene oxide synthase (LeAOS3, CYP74C3) was studied. Incubations of linoleic acid (9S)-hydroperoxide with dilute suspensions of LeAOS3 (10-20 s, 0 °C) yield mostly the expected allene oxide (12Z)-9,10-epoxy-10,12-octadecadienoic acid (9,10-EOD), which was detected as its methanol-trapping product. In contrast, the relative yield of 9,10-EOD progressively decreased when the incubations were performed with fourfold, tenfold, or 80-fold larger amounts of LeAOS3, while -ketol and the cyclopentenone rac-cis-10-oxo-11-phytoenoic acid (10-oxo-PEA) became the predominant products. Both the -ketol and 10-oxo-PEA were also produced when LeAOS3 was exposed to preformed 9,10-EOD, which was generated by maize allene oxide synthase (CYP74A). LeAOS3 also converted linoleic acid (13S)-hydroperoxide into the corresponding allene oxide, but with about tenfold lower yield of cyclopentenone. The results indicate that in contrast to the ordinary allene oxide synthases (CYP74A subfamily), LeAOS3 (CYP74C subfamily) is a multifunctional enzyme, catalyzing not only the synthesis, but also the hydrolysis and cyclization of allene oxide.

Keywords: allene oxide synthase • enzyme catalysis • metabolism • oxylipins • tomato

Source: ChemBioChem (2008) Vol. 9, Issue 15, p. 2498 - 2505

Determinants governing the CYP74 catalysis: Conversion of allene oxide synthase into hydroperoxide lyase by site-directed mutagenesis

2009-08-14T16:30:00.009+09:00

Bioinformatics analyses enabled us to identify the hypothetical determinants of catalysis by CYP74 family enzymes. To examine their recognition, two mutant forms F295I and S297A of tomato allene oxide synthase LeAOS3 (CYP74C3) were prepared by site-directed mutagenesis. Both mutations dramatically altered the enzyme catalysis. Both mutant forms possessed the activity of hydroperoxide lyase, while the allene oxide synthase activity was either not detectable (F295I) or significantly reduced (S297A) compared to the wild-type LeAOS3. Thus, both sites 295 and 297 localized within the “I-helix central domain” (“oxygen binding domain”) are the primary determinants of CYP74 type of catalysis.

Keywords: Cytochrome P450; CYP74 family; Allene oxide synthase; Hydroperoxide lyase; Site-directed mutagenesis

Source: FEBS Letters (2008) Vol. 582, Issues 23-24, Pages 3423-3428

My Article on BKCS 2009

2009-04-28T15:31:00.017+09:00

A Substrate Serves as a Hydrogen Atom Donor in the Enzyme-Initiated Catalytic Mechanism of Dual Positional Specific Maize Lipoxygenase-1

The maize lipoxgyenase-1 is a non-traditional dual positional specific enzyme and the reaction proceeds via enzyme-initiated catalysis. Bioinformatic analysis indicated that the maize lipoxygenase-1 is structurally more similar to soybean LOX1 than pea LOXN2 in that it has an additional external loop (residues 318-351) in the carboxy-terminal catalytic domain. We analyzed the dependence of product distribution on concentration of linoleic acid and monitored the formation of hydroperoxyoctadecadienoic acid as a function of enzyme concentration. Product distribution was strongly influenced by substrate concentration, such that kinetically-controlled regioisomers were enriched and thermodynamically-controlled regioisomers were depleted at high substrate concentration. Kinetic studies indicated that the formation of hydroperoxyoctadecadienoic acid saturated rapidly in an enzyme concentration-dependent manner, which implied that reactivation by reoxidation of inactive Fe(II) failed to occur. Our results support the previously proposed enzyme-initiated catalytic mechanism of the maize lipoxgyenase-1 and reveals that a substrate molecule serves as a hydrogen atom donor in its enzyme-initiated catalysis.

Bulletin of the Korean Chemical Society. (2009) vol 30, pages 719-723

[ Full Text in PDF ]

Biosynthesis and Metabolism of Jasmonates

2009-04-26T15:34:00.016+09:00

Jasmonates are derived from oxygenated fatty acids via the octadecanoid pathway and characterized by a pentacyclic ring structure. They have regulatory function as signaling molecules in plant development and adaptation to environmental stress. Until recently, it was the cyclopentanone jasmonic acid (JA) that attracted most attention as a plant growth regulator. It becomes increasingly clear, however, that biological activity is not limited to JA but extends to, and may even differ between its many metabolites and conjugates as well as its cyclopentenone precursors. The enzymes of jasmonate biosynthesis and metabolism may thus have a regulatory function in controlling the activity and relative levels of different signaling molecules. Such a function is supported by both the characterization of loss of function mutants in Arabidopsis, and the biochemical characterization of the enzymes themselves.

Source: Journal of Plant Growth Regulation (2005) vol. 23, p. 179-199

Jasmonate-Responsive Gene Expression

2009-03-21T23:50:00.002+09:00

Jasmonic acid (JA) and its volatile methyl ester (MeJA) belong to a family of lipid-derived signalling molecules that affect many aspects of plant life, including defence against certain pathogens and insects and some developmental processes. JA signal transduction leads to modulation of the expression of primary response genes, the products of which lead to the expression of secondary response genes. The ORCA3 transcription factor from Catharanthus roseus is a good candidate for a terminal component of the JA signal transduction pathway. To our knowledge, not a single component of the primary JA signal transduction pathway has been characterized to date in Arabidopsis. Many transcriptional components of secondary JA response pathways have been described in this model plant species, and are reviewed here. Our review advocates a strong adherence to signal transduction terminology as employed in the animal research field and in molecular biology textbooks, to simplify and correct current models about JA signal transduction leading to gene expression.

Source: Journal of Plant Growth Regulation (2005) Vol. 23, p. 200-210

12 Steps to Living Well

2009-02-04T19:25:00.004+09:00

Kevin Bungert is the owner, head coach and motivation behind Living Well Systems, Inc., a personal motivation and coaching company located in East Northport, New York, USA. Over the years he has developed 12 steps that he use in his coaching business to help people improve the problem areas of their lives. These 12 ideas are:

1. Define Your Purpose: What is it that you really want to accomplish in life, in your business, with regards to your health, with regards to your family and friends?

2. Be Honest With Yourself: Stop blaming someone else for your circumstances, your choices equals your life! Be honest with yourself! Because when you are, than you take back total control of your life.

3. Define Your Short: Everybody has goals, whether you write them down or not, you have goals. For example, without writing it down every month, you want to pay your mortgage, right? You want to eat healthy, right? You want to be in a loving relationship, right? You want some fun in your life, right?

4. Stay Positive and Believe In Yourself: Just believe in yourself and listen to the ACHIEVERS, not the whiners!

5. Learn New Information: The reason we must learn new information is because the knowl-edge we have right now has only gotten us to the level of production that we are at right now. We want to learn how to do more business!

6. Make Different Choices: Your business can change dramatically if you choose to make it happen!

7. Take Action: We must take action! That's where the great stuff is!

8. Reprogram Your Negative: Have you ever called yourself FAT, LAZY, STUPID, SILLY, UNORGANIZED, OLD, DISGUSTING, OR A LOSER? Well, that's negative self talk! And we have ALL done it, to some degree or other. The sad part is, it's so damaging. We call ourselves names all the time, that if other people said it, we would be really hurt! Well as adults, it is up to us to STOP OUR NEGATIVE SELF TALK AND RE-PLACE IT WITH MORE HELPFUL THOUGHTS!

9. Make New Associations: It means meeting new people who can help you achieve your goals. But the rub is, they don't come and knock on your door! We have to step out of our comfort zones and go seek out their help. Are you going to ask for help to make your life better?

10. Refine - Continually: Step #10, basically means put all 9 earlier steps into action and then take a look. If you want to make a change in a certain area of your life that is caus-ing you pain, begin to put these ideas into force in your life and keep try-ing! NOTHING CHANGES OVERNIGHT, PROGRESS TAKES TIME! You job is to NOT GIVE UP! EVER!

11. Help Others: Success in life is in what you do for others.

12. Don’t Be Afraid: We are all afraid that we are not good enough ins some way… We have to accept that some things are going to scare us… and we have to move forward anyway. START TODAY! DON'T BE AFRAID!

Food processing a tool to pesticide residue dissipation

2009-02-03T18:41:00.006+09:00

Food safety is an area of growing worldwide concern on account of its direct bearing on human health. The presence of harmful pesticide residues in food has caused a great concern among the consumers. Hence, world over to tackle food safety issues, organic farming is being propagated. However, due to several reasons, diffusion and acceptance of this approach in developing countries has been very slow. Therefore, it is important in the transient phase that some pragmatic solution should be developed to tackle this situation of food safety. Food processing treatments such as washing, peeling, canning or cooking lead to a significant reduction of pesticide residues. In this background this paper reviews the common food processing operations along with the degree of residue removal in each process. The processes reviewed include: baking, bread making, dairy product manufacture, drying, thermal processing, fermentation, freezing, infusion, juicing, malting, milling, parboiling, peeling, peeling and cooking, storage, storage and milling, washing, washing and cooking, washing and drying, washing and peeling, washing peeling and juicing and wine making. Extensive literature review demonstrates that in most cases processing leads to large reductions in residue levels in the prepared food, particularly through washing, peeling and cooking operations.

Source: Food Research International (2009) vol. 42, p. 26-40

Sleep! Sleep! Sleep!

2009-01-07T12:54:00.008+09:00

The following can interfere with sleep:
• Caffeine - coffee, sodas, tea, chocolate
• Tobacco
• A room that is too hot or too cold
• Light
• Noise
• An uncomfortable bed
• Using alcohol before bedtime
• Being hungry
• Eating a large meal close to bedtime
• A snoring bed partner
• A pet in the bedroom
• TV in the bedroom
• Getting too stirred up before bedtime can make it hard to go to sleep

Some tips that promote sleep:
• Make 8 hours of sleep a regular habit. Sleeping less during the week and trying to catch up on the weekend doesn’t work.
• Try to go to bed at the same time every night.
• If you have a clock that is always lit up, turn it so you can’t see the time.
• Exercise every day.
• Turn off your TV and computer an hour or two before bedtime.
• If you nap, keep it short and early in the day.
• Try reading before bedtime, but use a low-watt bulb.
• Do not eat a few hours before bedtime but don’t go to bed hungry. If you eat something, choose food that is light and nutritious. Avoid spicy or greasy food.
• Take a hot bath before retiring.
• If you feel you need to worry, tell yourself that you will only worry in the daytime. Make your bedroom a worryfree zone. Learn relaxation techniques to reduce stress and worrying.
• Listen to relaxation tapes before retiring.
• Do not lay awake in bed for more than 20 to 30 minutes. Get up, do something boring for a little while, and then go back to bed.
• Your bed is for sleep and sex. If you are not doing either of these, stay out of bed.

Cell signaling under salt, water and cold stresses

2009-01-05T14:07:00.007+09:00

Low temperature, drought, and high salinity are common stress conditions that adversely affect plant growth and crop production. The cellular and molecular responses of plants to environmental stress have been studied intensively (Thomashow, 1999; Hasegawa et al., 2000). Understanding the mechanisms by which plants perceive environmental signals and transmit the signals to cellular machinery to activateadaptive responses is of fundamental importance to biology. Knowledge about stress signal transduction is also vital for continued development of rational breeding and transgenic strategies to improve stress tolerance in crops. In this review, we first consider common characteristics of stress signal transduction in plants, and then examine some recent studies on the functional analysis of signaling components. Finally, we attempt to put these components and pathways into signal transduction networks that are grouped into three generalized signaling types.

Source: The Plant Cell (2002) vol. 14, p. S165-S183

5 Essential Weight Loss Foods

2008-12-24T13:41:00.002+09:00

According to Dr. Mao's Secrets of Longevity, there are five foods that will help you restore your body's ability to use energy and help you become your healthy weight.

1. Millet

A well-balanced diet should consist of whole grains instead of refined grains like white rice and pasta, and millet is a beneficial and delicious staple of this category of food. This non-glutinous grain is over 10-percent protein, has high amounts of fiber and B-complex vitamins, and because it isn't an acid forming food, is easy to digest.

2. Asparagus

When losing weight, it's important to favor chlorophyll-rich foods, including asparagus. Asparagus is a nutrient-rich vegetable packed with folate, vitamins A, C, and K, and fiber. Asparagus also contains a carbohydrate known as inulin (not to be confused with insulin) that promotes healthy bacteria in the large intestine - which in turn promotes a healthier digestive function.

3. Pomegranates

Eating a balanced diet to lose weight should include eating fresh fruits, and pomegranates are a wonderful example of a healthy, nutritious fruit that has antioxidant properties and will help prevent cancer. While the benefits of drinking pomegranate juice have gained a lot of attention recently, you will be more likely to lose weight by eating the fruit fresh to increase your fiber intake and keep the calories down.

4. Pine Nuts

Pine nuts are the edible seeds of pine trees and are considered an essential ingredient in the tasty Italian mixture pesto. Chinese medicine uses pine nuts to improve gastrointestinal tract and digestive functions, and pine nut oil is even used for appetite suppression. Pine nuts and other nuts are a tasty part of a well-balanced diet intended for weight loss.

5. Green Tea

It has been found that consuming large amounts of coffee and caffeine can lead to food cravings, increase one's appetite, and induce stress-related eating. Green tea is a wonderful alternative to coffee in that it does provide a little caffeine but also contains beneficial antioxidants. So drink up!

Source: health.yahoo.com

Studies on improved Agrobacterium-mediated transformation in two indica rice (Oryza sativa L.)

2008-12-22T10:31:00.010+09:00

Agrobacterium tumefaciens strain EHA 105 carrying binary vector pCAMBIA 1301 was used for transformation in two economically important highly recalcitrant indica rice cultivars HKR-46 and HKR-126. High concentrations of acetosyringone in the agrobacterium culture and co-cultivation medium proved to be indispensable for successful transformation. Embryogenic scutellar calli were used for transformation studies. Binary vector pCAMBIA 1301 have been proved efficient for transformation. The percent transient GUS expression found to be higher in cutivar HKR-126 (44.4%) as compared to HKR-46 (28.9%). The percent recovery of hygromycin resistant calli after 4-6 weeks on selection medium was maximum in HKR-126 (52.6%).

Source: African Journal of Biotechnology (2004) vol. 3, p. 572-575

Agrobacterium tumefaciens-Mediated Transformation of Maize Embryos Using a Standard Binary Vector System

2008-12-15T23:34:00.002+09:00

We have achieved routine transformation of maize (Zea mays) using an Agrobacterium tumefaciens standard binary (non-super binary) vector system. Immature zygotic embryos of the hybrid line Hi II were infected with A. tumefaciens strain EHA101 harboring a standard binary vector and cocultivated in the presence of 400 mg L1 L-cysteine. Inclusion of L-cysteine in cocultivation medium lead to an improvement in transient -glucuronidase expression observed in targeted cells and a significant increase in stable transformation efficiency, but was associated with a decrease in embryo response after cocultivation. The average stable transformation efficiency (no. of bialaphos-resistant events recovered per 100 embryos infected) of the present protocol was 5.5%. Southern-blot and progeny analyses confirmed the integration, expression, and inheritance of the bar and gus transgenes in R0, R1, and R2 generations of transgenic events. To our knowledge, this represents the first report in which fertile, stable transgenic maize has been routinely produced using an A. tumefaciens standard binary vector system.

Source: Plant Physiol. (2002) vol. 129, p. 13–22

Tamarind

2008-12-01T16:59:00.008+09:00

Common Name: Tamarind
Genus Species: Tamarindus indica
Family: Fabaceae
Origin: East Africa
Cultivated: India, Southeast Asian, West Indies

In India, Thailand, Cambodia, Southern Europe and Latin America, tamarind is a popular cooking and condiment flavor. The tree originated in Africa, but now it is cultivated mainly in South India and in the Caribbean.

A flavouring agent made from the leaf and fruit of the tamarind tree. The fruit is shaped like a long bean, inside which is a sour pulp. The pulp can be pressed to form a 'cake' or processed to make a paste. Small pieces of tamarind cake can be broken off and infused to create an acidic liquid flavouring used in Asian and Caribbean cooking. The sour fruit pulp contains tartaric acid; its taste goes well with meat and vegetable dishes, and it is used in marinades, vindaloos, curries, chutneys and Worcestershire sauce.

Tamarind is also used in drinks and in refreshing confectionary preparations. Tamarind juice is also available and some Asian supermarkets may sell tamarind pods, which can be eaten raw.

An improved Agrobacterium -mediated transformation protocol for recalcitrant elite indica rice cultivars

2008-11-21T23:31:00.002+09:00

We report here a high-efficiency transformation protocol for recalcitrant indica rice cultivars IR64 and IR72 with the selectable marker genehph and thegusA reporter gene. Factors that favor high-efficiency transformation were found to be use of 2-month-old mature seed-derived embryogenic calli, maltose as a source of carbon, a higher concentration of 2,4-dichlorophenoxyacetic acid, and both phytagel and agar as gelling agents. The putative transgenic (T0) plants were analyzed for integration of the transgene through polymerase chain reaction and Southern blotting analyses. Various factors thought to be responsible for increased transformation efficiency are discussed.

Source: Plant Molecular Biology Reporter (2005) vol. 23, p. 67-73

Agrobacterium -mediated transformation of the genome-sequenced poplar clone, nisqually-1 ( Populus trichocarpa )

2008-11-18T16:56:00.010+09:00

The US Department of Energy recently released a 6.8X draft of the genome sequence for Nisqually-1, a genotype of black cottonwood (Populus trichocarpa). To improve its utility for functional genomics research, having an efficient means for transformation and regeneration is necessary. To examine several parameters known to affect the transformation rate, we cocultivated leaf disc and stem explants with a strain ofAgrobacterium tumefaciens harboring a binary plasmid vector containing genes for both neomycin phosphotransferase (NPTII) and β-glucuronidase (GUS). Shoot regeneration from stem explants was observed in the presence of kanamycin when thidiazuron was incorporated in the selection medium. Transformation efficiency was influenced by the level of thidiazuron to which explants were exposed during the early stages of shoot induction. Histochemical assays revealed expression of theGUS gene in leaf, stem, and root tissues of transgenic plants. Polymerase chain reaction confirmed the presence of both selectable marker and reporter genes in all lines that stained positive for β-glucuronidase activity. By use of our modified protocol, transgenic plants were recovered within 6 mo at an efficiency of 6%, adequate to produce a large number of transgenic events with modest effort.

Source: Plant Molecular Biology Reporter (2004) vol. 22, p. 1-9.

Investigation of substrate binding and product stereochemistry issues in two linoleate 9-lipoxygenases

2008-10-14T23:57:00.002+09:00

Herein we characterize the Arabidopsis thaliana AtLOX1 and tomato (Solanum lycopersicum) LOXA proteins as linoleate 9S-lipoxygenases (9-LOX), and use the enzymes to test a model that predicts a relationship between substrate binding orientation and product stereochemistry. The cDNAs were heterologously expressed in E. coli and the proteins partially purified by nickel affinity chromatography using a N-terminal (His)(6)-tag. Both enzymes oxygenated linoleic acid almost exclusively to the 9S-hydroperoxide with turnover numbers of 300-400/s. AtLOX1 showed a broad range of activity over the range pH 5-9 (optimal at pH 6); tomato LOXA also showed optimal activity around pH 5-7 dropping off more sharply at pH 9. Site-directed mutagenesis of a conserved active site Ala (Ala562 in AtLOX1, Ala 564 in tomato LOXA, and typically conserved as Ala in S-LOX and Gly in R-LOX), revealed that substitution with Gly led to the production of a mixture of 9S- and 13R-hydroperoxyoctadecadienoic acids from linoleic acid. To follow up on earlier reports of 9-LOX metabolism of anandamide (van Zadelhoff et al. Biochem. Biophys. Res. Commun. 248:33-38, 1998), we also tested this substrate with the mutants, which produced predictable shifts in product profile, including a shift from the prominent 11S-hydroperoxy derivative of wild-type to include the 15R-hydroperoxide. These results conform to a model that predicts a head-first substrate binding orientation for 9S-LOX. We also found that linoleoyl-phosphatidylcholine is not a 9S-LOX substrate, which is consistent with this conclusion.

Source: Lipids (2008) vol. 43, p. 979-987

Agrobacterium-mediated transformation of an elite indica rice for insect resistance

2008-10-10T23:27:00.002+09:00

In India, rice occupies the largest area among all crops and accounts for as much as 21 % of the total cropped area. Rice plants are prone to attack from many pests and pathogens. Among them coleopteran pests are well known for causing extensive damage. Sitophilus oryzae causes severe damage to rice seeds during storage. Alpha amylase inhibitor gene isolated from Phaseolus vulgaris seeds was introduced into Basmati rice (PB1) through Agrobacterium -mediated transformation. A total of 174 hygromycin-resistant plants were regenerated. Most of these plants were GUS-positive. PCR analysis and Southern hybridization confirmed the presence of 4.9 kb alpha amylase inhibitor gene in transformed plants. Western blot confirmed presence of alpha amylase inhibitor protein. Results of the bioassay study revealed significant reduction in survival rate of rice weevil, S. oryzae reared on transgenic rice seeds.

Source: Current Science, Vol 90 (2006) Page 829-835

Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA

2008-10-01T12:34:00.012+09:00

A large number of morphologically normal, fertile, transgenic rice plants were obtained by co-cultivation of rice tissues with Agrobacterium tumefaciens. The efficiency of transformation was similar to that obtained by the methods used routinely for transformation of dicotyledons with the bacterium. Stable integration, expression and inheritance of transgenes were demonstrated by molecular and genetic analysis of transformants in the R0, R1 and R2 generations. Sequence analysis revealed that the boundaries of the T-DNA in transgenic rice plants were essentially identical to those in transgenic dicotyledons. Calli induced from scutella were very good starting materials. A strain of A. tumefaciens that carried a so-called 'super-binary' vector gave especially high frequencies of transformation of various cultivars of japonica rice that included Koshihikari, which normally shows poor responses in tissue culture.

Source: The Plant Journal (1994) vol. 6, p. 271-282

Benefits of Biotechnology

2008-09-22T16:31:00.004+09:00

• Protection of the environment: Scientists have made some foods, such as papayas and potatoes, more resistant to disease. These crops need less chemical spray to protect them from harmful insects or viruses, which is better for water and wildlife. Other crops are protected from herbicides that are used to control weeds, thus allowing farmers to conserve soil by tilling the ground less often.

• Greater Crop Yields: Farmers can use biotechnology to help plants survive, warding off insects and better tolerance to herbicides. This allows a better harvest from these hardier plants.

• Better Tasting, Fresher Foods: Sweeter peppers and tomatoes that ripen more slowly are examples of how biotechnology can produce fresher and better tasting food.

• Grow more food on less land: By the year 2050, the earth’s population is estimated to be nine billion people. Using biotechnology, farmers can produce more crops on the land they already have. This way, countries do not have to devote more land to farming. In turn, developing countries can benefit most, since they will have the largest population growth.

• Keep food safe to eat: Scientists can more accurately find unwanted viruses and bacteria that may be present in food. This will cause an even lower risk of food-borne illnesses. Some types of fungus, which can be found in corn, release substances that can harm animals that eat them. These substances are already regulated in the United States, and biotechnology provides another tool that can help further reduce the amount of these substances in corn.

• New food varieties: Biotechnology can extend advances in cross-breeding, allowing for new food varieties. For example, seedless melons and mini avocadoes. Farmers can also develop food with better flavor and a better nutrient profile.

The Lipoxygenase pathway

2008-09-15T23:15:00.002+09:00

Lipid peroxidation is common to all biological systems, both appearing in developmentally and environmentally regulated processes of plants. The hydroperoxy polyunsaturated fatty acids, synthesized by the action of various highly specialized forms of lipoxygenases, are substrates of at least seven different enzyme families. Signaling compounds such as jasmonates, antimicrobial and antifungal compounds such as leaf aldehydes or divinyl ethers, and a plant-specific blend of volatiles including leaf alcohols are among the numerous products. Cloning of many lipoxygenases and other key enzymes within the lipoxygenase pathway, as well as analyses by reverse genetic and metabolic profiling, revealed new reactions and the first hints of enzyme mechanisms, multiple functions, and regulation. These aspects are reviewed with respect to activation of this pathway as an initial step in the interaction of plants with pathogens, insects, or abiotic stress and at distinct stages of development.

Source: Annual review of plant biology (2002) vol. 53, p. 275-297

The Jasmonate Signal Pathway

2008-09-10T12:05:00.006+09:00

Plant responses to many biotic and abiotic stresses are orchestrated locally and systemically by signaling molecules known as the jasmonates (JAs). JAs also regulate such diverse processes as pollen maturation and wound responses in Arabidopsis.
The JA signal pathway involves several signal transduction events: the perception of the primary wound or stress stimulus and transduction of the signal locally and systemically; the perception of this signal and induction of JA biosynthesis; the perception of JA and induction of responses; and finally, integration of JA signaling with outputs from the SA, ethylene, and other signaling pathways.

Source: The Plant Cell (2002) vol. 14, p. S153-S164

Which Fruits and Vegetables Are the Most Toxic?

2008-09-09T23:24:00.010+09:00

10 most toxic conventionally grown fruits and vegetables (in order of pesticide toxicity levels):

- Peaches
- Apples
- Sweet Bell Peppers (11 different pesticides found on a single pepper)
- Celery
- Nectarines
- Strawberries
- Cherries
- Lettuce
- Grapes
- Pears (86 percent of samples tested with detectable pesticides)

10 least toxic conventionally grown fruits and vegetables (in descending order of pesticide toxicity levels):

- Cabbage
- Bananas
- Kiwi
- Asparagus (7 percent of samples tested with detectable pesticides)
- Sweet Peas
- Mangoes
- Pineapples
- Sweet Corn
- Avocado
- Onions (1 pesticide found on a single onion)

Source: Health.msn.com

Lipoxygenase biocatalysis: a survey of asymmetric oxygenation

2008-08-27T15:03:00.011+09:00

Lipoxygenases are a group of non-heme iron containing dioxygenases catalyzing the addition of molecular oxygen to poly unsaturated fatty acids in a stereospecific as well as regiospecific way and involved in the biosynthesis of inflammatory mediators, in cell differentiation and atherogenesis. Synthetic use of lipoxygenase over the last decade for total synthesis of several natural products, e.g. 6(R)-lopoxin A, 5(S)-HPETE, (R) and (S) coriolide is discussed in this review. In the second part of the review we have discussed on the LOX biocatalysis of unnatural substrates mimicking natural substrates, lipoxygenase biocatalysis in non-conventional media, the use of immobilized LOX and biocatalysis with engineered lipoxygenase. The factors governing the regiospecificity as well as stereospecificity of LOX biocatalysis will also be discussed here. From the synthetic chemist’s point of view, the asymmetric oxygenation reaction of unnatural substrates has a tremendous potential. The products of these reactions are important chiral building blocks for the total synthesis of numerous biological active natural products.

Source: Journal of Molecular Catalysis B: Enzymatic (2003) vol. 26, p. 3-28

A New 9-Lipoxygenase cDNA from Developing Rice Seeds

2008-08-21T23:11:00.002+09:00

We isolated a novel C9 position specific lipoxygenase (r9-LOX1) cDNA from developing rice seeds. The enzymatic features of r9-LOX1 resembled those of rice LOX-L3 known to be contained in rice germ and to have C9-specific LOX activity. However, the expression level of the r9-LOX1 gene was higher in imbibed seeds rather than developing seeds. A homology search against the rice nucleotide database revealed the r9-LOX1 gene to be on rice chromosome 3 (accession number AC093017). The restriction enzyme map of the reported genomic sequence agreed with the result of the Southern blot analysis for the r9-LOX1. The enzyme could be useful for in vitro synthesis of 9,10-ketol-octadecadienoic acid.

Source: Plant and Cell Physiology (2003) vol. 44, p. 1168-1175

FeatuDefects in Allene Oxide Synthase and 12-Oxa-Phytodienoic Acid Reductase Alter the Resistance to Pseudomonas syringae and Botrytis cinerea

2008-07-31T11:04:00.012+09:00

Jasmonic acid and its methyl ester are signalling molecules involved in regulating development and stress responses in plants. 12-Oxo-phytodienoic acid, a precursor in jasmonic acid biosynthesis, is also biologically active. Both oxylipins accumulate after pathogen infection. To understand the function of 12-oxo-phytodienoic acid vs. jasmonic acid in plant pathogen interactions, the susceptibility of the dde2 mutant, defective in 12-oxo-phytodienoic acid and jasmonic acid biosynthesis and the opr3 mutant, which synthesizes 12-oxo-phytodienoic acid but not jasmonic acid was investigated. dde2 and opr3 were more resistant to Pseudomonas syringae. The effect was stronger in the dde2 mutant as in opr3 indicating that both oxylipins regulate pathogen resistance to P. syringae. dde2 was more susceptible to Botrytis cinerea suggesting that 12-oxo-phytodienoic acid is important in mediating the defence against B. cinerea. Expression of Pr1 and Pr2 was increased in both mutants after mock infiltration but not or only slightly enhanced after P. syringae infection.

Source: Journal of Phytopathology (2006) vol. 154 , p. 740 - 744

Recent developments in biochemistry of the plant lipoxygenase pathway

2008-07-21T22:59:00.002+09:00

Lipoxygenase (LOX, EC 1.13.11.12) is a dioxygenase enzyme widespread in aerobic organisms, including plants. The enzyme oxygenates methylene interrupted polyenoic fatty acids, converting them into hydroperoxides. The predominant substrates of LOXs in plants are linoleic and linolenic acids. Specificities of LOX action and availability of enzymes of hydroperoxide metabolism detemine the diversity of the LOX pathway products. Increasing evidence in the literature demonstrates the great physiological importantance of the plant LOX pathway products.

Source: Progress in Lipid Research (1998) vol. 37 , p. 317-352

Divinyl ether synthesis in garlic bulbs

2008-07-19T21:45:00.002+09:00

Formation of 13-lipoxygenase-derived divinyl ethers has been described in garlic bulbs. Here, the identification of a cDNA from garlic is described, which encodes for an enzyme that corresponds to divinyl ether synthases (DES). The recombinant protein was expressed in Escherichia coli and shown to metabolize 13-hydroperoxy as well as 9-hydroperoxy linole(n)ic acid to etherole(n)ic and colnele(n)ic acid, respectively. This biochemical feature classifies it as a member of the CYP74C subfamily of cytochrome P-450 enzymes. Product analysis after incubation of purified recombinant enzyme and fatty acid hydroperoxides revealed the formation of a mixture of different cis/trans isomers with one isomer often dominant. RNA blot analyses showed a constitutive expression of DES transcripts predominant in below-ground organs of garlic. By exogenous application of salicylic acid and sorbitol, but not by methyljasmonate, the transcript was also induced in leaves. Whereas the prominent divinyl ether in garlic was the 13-lipoxygenase-derived etheroleic acid, analysis of transgenic Arabidopsis expressing garlic DES showed that 9-lipoxygenase-derived colnelenic acid dominated 24 h after wounding. These data indicate that the product pattern of this DES from garlic depends on the substrate availability and that the enzyme is the first member in the group of 9/13-DES.

Source: J Exp Bot. (2008) vol. 59, p. 907-915

Animals and Plants as National Symbols of Cambodia

2008-07-06T01:58:00.009+09:00

1. Kouprey (Bos sauveli) is the National Mammal of Cambodia
2. Giant Ibis (Pseudibis gigantea) is the National Bird of Cambodia
3. Royal Turtle (Batagur baska) is the National Reptile of Cambodia
4. Giant Mekong Barb (Catlocarpio siamensis) is the National Fish of Cambodia
5. Sugar Palm (Borassus flabellifer) is the National Tree of Cambodia
6. Rumdul (Mitrella mesnyi) is the National Flower of of Cambodia
7. Chicken Egg Banana (Musa aromatica) is the National Fruit of Cambodia

[PDF Full text]

Adapted from Forestry Administration of Cambodia

Best fuel for your workout

2008-07-03T14:58:00.010+09:00

Good food choices to fuel your workout include high-carbohydrate, low-fat snacks such as:

- a banana with yogurt;

- a whole-wheat toasted bagel with fruit spread; or

- a quick bowl of instant oatmeal.

After your workout, it’s essential to refuel your body with a carbohydrate-protein blend, such as:

- a peanut butter sandwich;

- a small handful of walnuts or almonds; or

- a fruit smoothie with some protein powder.

Don't forget to drink water before, during and after your workout to avoid dehydration.

7 Foods to Keep You Young

2008-06-30T11:22:00.011+09:00

There are many foods that you are eating everyday. Seven foods here can keep you younger and long life are Olive oil, Yogurt, Fish, Chocolate, Nuts, Wine, and Blueberries.

Source: Health.msn.com

Jasmonates as signals in the wound response

2008-06-15T22:54:00.002+09:00

Plant responses to wounding and herbivore attack are orchestrated by complex signaling pathways that link the production of chemical and physical signals at the wound site to activation of gene expression and other cellular processes. The systemic nature of many wound-induced responses provides an attractive opportunity to study intercellular signaling pathways that operate over long distances within the plant. Genetic dissection of the wound-response pathway in tomato indicates that (1) systemin and its precursor protein, prosystemin, are upstream components of an intercellular signaling cascade that requires the biosynthesis and action of jasmonic acid (JA); and (2) physiological processes regulated by this pathway confer host resistance to a broad spectrum of plant invaders. Grafting experiments conducted with mutants defective in systemic wound signaling indicate that systemin functions at or near the wound site to trigger the production of JA, which in turn acts non-cell autonomously to promote systemic defense responses. The location of JA biosynthetic enzymes within the companion cell-sieve element complex of vascular bundles, together with the accumulation of JA in vascular tissues, support a role for jasmonates as phloem-mobile signals. The recent discovery of enzymes involved in the metabolism of JA to volatile methyl-JA and bioactive JA-amino acid conjugates has potential implications for the mechanism by which JA promotes wound signaling. Species-specific differences in the mechanism of wound signaling appear to reflect the way in which the wound-induced jasmonate pathway is regulated by other signals including systemin, cell wall-derived oligosaccharides, ethylene, and insect-derived elicitors. Adding to the complexity of the wound-induced jasmonate cascade are wound-signaling pathways that operate independently of JA.

Source: Journal of Plant Growth Regulation (2004) vol. 23 , p. 223-237

Oxylipin metabolism in response to stress

2008-06-10T00:38:00.009+09:00

Oxylipins comprise a group of biologically active compounds whose structural diversity is generated by the coordinate action of lipases, lipoxygenases, and a group of cytochromes P450 that are specialized for the metabolism of hydroperoxy fatty acids. Research on oxylipins has focused mainly on the biosynthesis of the plant signaling molecule jasmonic acid, and its role in the regulation of developmental and defense-related processes. Recent genetic studies indicate that metabolic precursors of jasmonate are active as signals in their own right, and that the synthesis and perception of jasmonates is critical for wound-induced systemic defense responses. Increasing evidence indicates that the collective biological importance of oxylipins in plants is comparable to that of the eicosanoid family of lipid mediators in animals.

Source: Current opinion in plant biology (2002) vol. 5, p. 230-236

From plant genomics to breeding practice

2008-05-21T22:51:00.002+09:00

New alleles are constantly accumulated during intentional cropselection. The molecular understanding of these alleles hasstimulated new genomic approaches to mapping quantitativetrait loci (QTL) and haplotype multiplicity of the genes concerned. A limited number of quantitative trait nucleotides responsible forQTL variation have been described, but an acceleration in theirrate of discovery is expected with the adoption of linkagedisequilibrium and candidate gene strategies for QTL finemapping and cloning. Additional layers of regulatory variationhave been studied that could also contribute to the molecularbasis of quantitative genetics of crop traits. Despite thisprogress, the role of marker-assisted selection in plant breedingwill ultimately depend on the genetic model underlyingquantitative variation.

Source: Current opinion in biotechnology (2003) vol. 14, p. 214-219

Detection of a pathway from linoleate to a novel cyclopentenone: cis-12-oxo-10-phytoenoic acid in sunflower roots

2008-05-17T19:50:00.016+09:00

The lipoxygenase pathway in sunflower roots was studied in vitro. A preliminary incubation of linoleic acid with 15 000 g supernatant of homogenate of sunflower roots (1.5-6 days after germination) revealed the predominant activity of 13-lipoxygenase. The exogenously added linoleic acid 13-hydroperoxide is further utilized through two competing pathways. One of them is directed towards formation of the ketodiene (9Z,11E)-13-oxooctadeca-9,11-dienoic acid. The second pathway, which is controlled by allene oxide synthase, leads to the formation of an alpha-ketol and a novel cyclopentenone, rac-cis-12-oxo-10-phytoenoic acid (12-oxo-PEA) via a short-lived allene oxide. Unexpectedly, the cyclopentenone 12-oxo-PEA is the predominant allene oxide synthase product. Identification of cis-12-oxo-PEA was confirmed by its UV, mass, (1)H NMR and 2D-COSY spectral data. The highest yield of 12-oxo-PEA is observed in very young roots (1.5-2 days after germination). The results of methanol-trapping experiments demonstrate that both 12-oxo-PEA and alpha-ketol are formed through the unstable allene oxide intermediate, (9Z)-12,13-epoxyoctadeca-9,11-dienoic acid, which is the primary product of allene oxide synthase. Since 12-oxo-PEA is a jasmonate congener, its biosynthesis in plants might be of physiological importance.

Source: Chembiochem. (2007) vol. 18, p. 2275-2280

Oxygenation of arachidonoyl lysophospholipids by lipoxygenases from soybean, porcine leukocyte, or rabbit reticulocyte

2008-05-10T20:42:00.002+09:00

Oxygenation of arachidonoyl lysophosphatidylcholine (lysoPC) or arachidonoyl lysophosphatidic acid (lysoPA) by lipoxygenase (LOX) was examined. The oxidized products were identified by HPLC/UV spectrophotometry/mass spectrometry analyses. Straight-phase and chiral-phase HPLC analyses indicated that soybean LOX-1 and rabbit reticulocyte LOX oxygenated arachidonoyl lysophospholipids mainly at C-15 with the S form as major enantiomer, whereas porcine leukocyte LOX oxygenated at C-12 with the S form. Next, the sequential exposure of arachidonoyl-lysoPC to soybean LOX-1 and porcine leukocyte LOX afforded two major isomers of dihydroxy derivatives with conjugated triene structure, suggesting that 15(S)-hydroperoxyeicosatetraenoyl derivatives were converted to 8,15(S)-dihydroxyeicosatetraenoyl derivatives. Separately, arachidonoyl-lysoPA, but not arachidonoyl-lysoPC, was found to be susceptible to double oxygenation by soybean LOX-1 to generate a dihydroperoxyeicosatetraenoyl derivative. Overall, arachidonoyl lysophospholipids were more efficient than arachidonic acid as LOX substrate. Moreover, the catalytic efficiency of arachidonoyl-lysoPC as substrate of three lipoxygenases was much greater than that of arachidonoyl-lysoPA or arachidonic acid. Taken together, it is proposed that arachidonoyl-lysoPC or arachidonoyl-lysoPA is efficiently oxygenated by plant or animal lipoxygenases, C12- or C15-specific, to generate oxidized products with conjugated diene or triene structure.

Source: J Agric Food Chem. (2008) vol. 56, p. 1224-1232

The New Superfruits

2008-05-01T11:34:00.008+09:00

For Perfect Blood Pressure
- Good: Bananas
- Better: Fresh figs

To Protect Your Heart and Fight Disease
- Good: Red grapes
- Better: Lychee

For Beautiful Skin
- Good: Orange
- Better: Guava

To Lower Cholesterol
- Good: Apples
- Better: Asian pears

To Fight Cancer
- Good: Watermelon
- Better: Papaya

Source: Health.msn.com

Basic Conversation in Korean

2008-04-30T10:08:00.013+09:00

GM crops on trial: Technological development as a real-world experiment

2008-04-29T00:35:00.012+09:00

Through the European controversy over agricultural biotechnology, genetically modified (GM) crops have been evaluated for an increasingly wide range of potential effects. As the experimental phase has been extended into commercial practices, the terms for product approval have become more negotiable and contentious. To analyse the regulatory conflicts, this paper links three theoretical perspectives: issue-framing, agri-environmental discourses, and technological development as a real-world experiment.

Agri-biotechnological risks have been framed by contending discourses, which attribute moral meanings to the agricultural environment. Agri-biotech proponents have emphasised eco-efficiency benefits, which can remedy past environmental damage, while critics have framed ‘uncontrollable risks’ in successively broader ways through ominous metaphors of environmental catastrophe. Regulatory authorities have translated those metaphors into measurable biophysical effects. They anticipate and design commercial use as a ‘real-world experiment’, by assigning greater moral-legal responsibility to agro-industrial operators who handle GM products.

Expert-regulatory debate reflexively considers the social discipline necessary to prevent harm, now more broadly defined than before. Official procedures undergo tensions between predicting, testing and prescribing operator behaviour. In effect, GM crops have been kept continuously ‘on trial’.

Source: Futures (2007) vol. 39, p. 408-431

Oxylipin Pathway in Rice and Arabidopsis

2008-04-21T22:46:00.002+09:00

Plants have evolved complex signaling pathways to coordinate responses to developmental and environmental information. The oxylipin pathway is one pivotal lipid-based signaling network, composed of several competing branch pathways, that determines the plant's ability to adapt to various stimuli. Activation of the oxylipin pathway induces the de novo synthesis of biologically active metabolites called "oxylipins". The relative levels of these metabolites are a distinct indicator of each plant species and determine the ability of plants to adapt to different stimuli. The two major branches of the oxylipin pathway, allene oxide synthase (AOS) and hydroperoxide lyase (HPL) are responsible for production of the signaling compounds, jasmonates and aldehydes respectively. Here, we compare and contrast the regulation of AOS and HPL branch pathways in rice and Arabidopsis as model monocotyledonous and dicotyledonous systems. These analyses provide new insights into the evolution of JAs and aldehydes signaling pathways, and the complex network of processes responsible for stress adaptations in monocots and dicots.

Source: Journal of Integrative Plant Biology (2007) vol. 49, p. 43-51

5 Foods That Feed Cholesterol

2008-04-14T10:32:00.010+09:00

5 Foods That Feed Cholesterol:

1. Whole-milk dairy products
2. Processed meats
3. Fast-food fries
4. Tropical oils
5. Baked goods

Source: Health.msn.com

Lipases for biotechnology

2008-03-21T22:43:00.002+09:00

Lipases constitute the most important group of biocatalysts for biotechnological applications. The high-level production of microbial lipases requires not only the efficient overexpression of the corresponding genes but also a detailed understanding of the molecular mechanisms governing their folding and secretion. The optimisation of industrially relevant lipase properties can be achieved by directed evolution. Furthermore, novel biotechnological applications have been successfully established using lipases for the synthesis of biopolymers and biodiesel, the production of enantiopure pharmaceuticals, agrochemicals, and flavour compounds.

Source: Current opinion in biotechnology, Vol 13, 2002, Pages 390-397

Rice octadecanoid pathway

2008-03-18T22:41:00.002+09:00

Plant jasmonic acid (JA) and structurally similar animal prostaglandins play pivotal roles in regulating cellular responses against environmental cues, including the innate immune response(s). In plants, JA and its immediate precursor 12-oxo-phytodienoic acid (OPDA) are synthesized by the octadecanoid pathway, which employs at least five enzymes (lipase, lipoxygenase, allene oxide synthase and cyclase, and OPDA reductase), in addition to the enzymes involved in the beta-oxidation steps. Genetic, molecular, and biochemical analyses have led to the identification of almost all the genes of the octadecanoid pathway in Arabidopsis--a model dicotyledonous plant. In this regard, rice (Oryza sativa L.)--an important socio-economic monocotyledonous model research plant--remains poorly characterized. Until now, no gene has been specifically associated with this pathway. It is therefore of utmost importance to identify, characterize, and assign the pathway specific genes in rice. In this review, we have surveyed the rice genome, extracted a large number of putative genes of the octadecanoid pathway, and discussed their relationship with the known pathway genes from other plant species. Moreover, the achievements made so far on the rice octadecanoid pathway have also been summarized to reflect the contribution of rice towards extending our knowledge on this critical pathway in plants.

Source: Biochemical and Biophysical Research Communications, Vol 317, 2004, Pages 1-15

Agrobacterium-mediated genetic transformation of plants: biology and biotechnology

2008-03-16T23:48:00.015+09:00

Agrobacterium-mediated genetic transformation is the dominant technology used for the production of genetically modified transgenic plants. Extensive research aimed at understanding and improving the molecular machinery of Agrobacterium responsible for the generation and transport of the bacterial DNA into the host cell has resulted in the establishment of many recombinant Agrobacterium strains, plasmids and technologies currently used for the successful transformation of numerous plant species. Unlike the role of bacterial proteins, the role of host factors in the transformation process has remained obscure for nearly a century of Agrobacterium research, and only recently have we begun to understand how Agrobacterium hijacks host factors and cellular processes during the transformation process. The identification of such factors and studies of these processes hold great promise for the future of plant biotechnology and plant genetic engineering, as they might help in the development of conceptually new techniques and approaches needed today to expand the host range of Agrobacterium and to control the transformation process and its outcome during the production of transgenic plants.

Source: Current Opinion in Biotechnology, Volume 17, 2006, Pages 147-154

5 Foods That Fight Cholesterol

2008-03-06T12:13:00.009+09:00

There’s no denying that a healthy diet is the first line of defense against rising cholesterol. “If you eat a predominantly plant-based diet—with lots of fruits and vegetables plus some fish—you are on the right track to keeping your cholesterol at a healthy level,” says Lisa Dorfman, a registered dietitian and spokeswoman for the American Dietetic Association. That said, certain so-called super-foods can actually help lower bad cholesterol and/or increase the good cholesterol. Ideally, you want to shoot for total cholesterol under 200, with LDL (the bad one) under 110 and HDL (the good one) greater than 35.

Try to incorporate more of these foods into your daily diet:

1. Almonds
2. Oatmeal
3. Fish
4. Red wine
5. Soy

Source: Health.msn.com

8 Foods You Should Eat Every Day

2008-03-03T19:20:00.014+09:00

We should eat 8 foods every day are
1. Spinach
2. Yogurt
3. Tomatoes
4. Carrots
5. Blueberries
6. Black Beans
7. Walnuts
8. Oats

Source: Health.msn.com

Transgenic Rice Breeding for Abiotic Stress Tolerance-Present and Future

2008-02-29T00:09:00.017+09:00

Environmental stresses and the continuing deterioration of arable land, along with an explosive increase in world population, pose serious threats to global agricultural production and food security. Improving the tolerance of the major crop plants to abiotic stresses has been a main goal in agriculture for a long time. As rice is considered one of the major crops, the development of new cultivars with enhanced abiotic stress-tolerance will undoubtedly have an important effect on global food production. The transgenic approach offers an attractive alternative to conventional techniques for the genetic improvement of rice cultivars. In recent years, an array of stress-related genes has already been transferred to rice to improve its resistance against abiotic stresses. Many transgenic rice plants with enhanced abiotic stress-tolerance have been obtained. This article focuses on the progress in the study of abiotic stress tolerance in transgenic rice breeding.

Source: Chinese Journal of Biotechnology, Vol 23, 2007, Pages 1-7

My article on Organic Letters 2007

2008-02-27T02:40:00.018+09:00

Regiochemical and stereochemical evidence for enzyme-initiated catalysis in dual positional specific maize lipoxygenase-1

Dual positional specific maize lipoxygenase-1 catalyzed the formation of racemic mixtures of four possible regioisomers and was strongly inhibited by the radical scavenger, 4-hydroxy-2,2,6,6-tetramethyl-1-piperidinoxy radical. Molecular modeling studies indicated that the oxygen-binding cavity is segregated from the substrate-binding cavity. The data suggest that a bis-allylic radical reaction intermediate is generated enzymatically, released from the enzyme active site, and subsequently oxygenated outside of the enzyme active site by a nonenzymatic mechanism.

[ Full Text in PDF ]

My article on J Biochem Mol Biol 2007

2008-02-27T02:35:00.015+09:00

Biochemical characterization of the dual positional specific maize lipoxygenase and the dependence of lagging and initial burst phenomenon on pH, substrate, and detergent during pre-steady state kinetics

The wound-inducible lipoxygenase obtained from maize is one of the nontraditional lipoxygenases that possess dual positional specificity. In this paper, we provide our results on the determination and comparison of the kinetic constants of the maize lipoxygenase, with or without detergents in the steady state, and characterization of the dependence of the kinetic lag phase or initial burst, on pH, substrate, and detergent in the pre-steady state of the lipoxygenase reaction. The oxidation of linoleic acid showed a typical lag phase in the pre-steady state of the lipoxygenase reaction at pH 7.5 in the presence of 0.25% Tween-20 detergent. The reciprocal correlation between the induction period and the enzyme level indicated that this lag phenomenon was attributable to the slow oxidative activation of Fe (II) to Fe (III) at the active site of the enzyme as observed in other lipoxygenase reactions. Contrary to the lagging phenomenon observed at pH 7.5 in the presence of Tween-20, a unique initial burst was observed at pH 6.2 in the absence of detergents. To our knowledge, the initial burst in the oxidation of linoleic acid at pH 6.2 is the first observation in the lipoxygenase reaction. Kinetic constants (K(m) and k(cat) values) were largely dependent on the presence of detergent. An inverse correlation of the initial burst period with enzyme levels and interpretations on kinetic constants suggested that the observed initial burst in the oxidation of linoleic acid could be due to the availability of free fatty acids as substrates for binding with the lipoxygenase enzyme.

[ Full Text in PDF ]

My article on Bioorganic Chemistry 2003

2008-02-27T01:39:00.019+09:00

A point mutation of valine-311 to methionine in Bacillus subtilis protoporphyrinogen oxidase does not greatly increase resistance to the diphenyl ether herbicide oxyfluorfen

In an effort to asses the effect of Val311Met point mutation of Bacillus subtilis protoporphyrinogen oxidase on the resistance to diphenyl ether herbicides, a Val311Met point mutant of B. subtilis protoporphyrinogen oxidase was prepared, heterologously expressed in Escherichia coli, and the purified recombinant Val311Met mutant protoporphyrinogen oxidase was kinetically characterized. The mutant protoporphyrinogen oxidase showed very similar kinetic patterns to wild type protoporphyrinogen oxidase, with slightly decreased activity dependent on pH and the concentrations of NaCl, Tween 20, and imidazole. When oxyfluorfen was used as a competitive inhibitor, the Val311Met mutant protoporphyrinogen oxidase showed an increased inhibition constant about 1.5 times that of wild type protoporphyrinogen oxidase. The marginal increase of the inhibition constant indicates that the Val311Met point mutation in B. subtilis protoporphyrinogen oxidase may not be an important determinant in the mechanism that protects protoporphyrinogen oxidase against diphenyl ether herbicides.

[ Full Text in PDF ]

A novel lipoxygenase gene from developing rice seeds confers dual position specificity and responds to wounding and insect attack

2008-01-26T21:39:00.002+09:00

OsLOX1 is a novel full-length cDNA isolated from developing rice seeds. We have examined its biochemical properties and expression patterns. The protein has dual positional specificity, as it releases both C-9 and C-13 oxidized products in a 4:3 ratio. OsLOX1 transcripts were detected at low abundance in immature seeds and newly germinated seedlings, but accumulate rapidly and transiently in response to wounding or brown planthopper (BPH) attack, reaching a peak 3 h after wounding and 6 h after insect feeding. We produced transgenic rice lines carrying either sense or antisense constructs under the control of a cauliflower mosaic virus 35S promoter, and these rice lines showed altered OsLOX1 activity. In all of the antisense lines and more than half of the sense lines the expression levels of OsLOX1, the levels of enzyme activity, and the levels of the endogenous OsLOX1 products (jasmonic acid, (Z)-3-hexenal and colneleic acid) at 6, 48, and 48 h after BPH feeding respectively, were below the levels found in non-transgenic control plants; yet, the levels in the remaining sense transformants were enhanced relative to controls. Transformants with a lower level of OsLOX1 expression were less able to tolerate BPH attack, while those with enhanced OsLOX1 expression were more resistant. Our data suggest that the OsLOX1 product is involved in tolerance of the rice plant to wounding and BPH attack.

Source:

Transgenic animal production and animal biotechnology

2008-01-23T22:25:00.002+09:00

Considerable progress has been made in methods for production of transgenic livestock; beginning with pronuclear microinjection over 20 years ago. New methods, including the use of viral vectors, sperm-mediated gene transfer and somatic cell cloning, have overcome many of the limitations of pronuclear microinjection. It is now possible to not only readily make simple insertional genetic modifications, but also to accomplish, more complex, homozygous gene targeting and artificial chromosome transfer in livestock.

Source: Theriogenology, Vol 67, 2007, Pages 127-133

Genes to feed the world: The weakest link?

2008-01-20T22:19:00.002+09:00

The new science of genomics has an important contribution to make to the development of agriculture in the ‘Third World’ and thus to improving the livelihoods of many of the poorest people on earth. The persistence of poverty demonstrates, however, that this contribution is not yet being made. We therefore examine the web of connections between the two and enquire whether they could be strengthened in some way, perhaps by the addition of some vital ‘missing link’, so that the contribution from genomics can be maximised. Since the use of molecular markers to help develop improved crop varieties (marker-assisted selection) constitutes one of the main ways in which genomic knowledge could be used to improve rural livelihoods, we review this technology and discuss the ways in which it could contribute to plant breeding. We argue that marker-assisted selection (MAS) is likely to be most effective when it is both (i) linked to the farmers who are expected to cultivate the new crop varieties that will eventually be developed; and (ii) integrated with plant breeding and with a range of complementary disciplines. Such integration necessarily involves the formation of effective trans-disciplinary teams, an undertaking that is often problematic, and so the final part of this article considers the processes that this involves and suggests ways in which they may be managed effectively.

Adapted from Food Policy, Vol 32, 2007, Pages 459-479

Identification of a functional allene oxide synthase-lipoxygenase fusion protein in the soft coral Gersemia fruticosa suggests the generality of ...

2007-12-06T22:37:00.002+09:00

Identification of a functional allene oxide synthase-lipoxygenase fusion protein in the soft coral Gersemia fruticosa suggests the generality of this pathway in octocorals

The conversion of fatty acid hydroperoxides to allene epoxides is catalysed by a cytochrome P450 in plants. In contrast, in the coral Plexaura homomalla, a catalase-related hemoprotein fused to the lipoxygenase (LOX) was found to function as an allene oxide synthase. This work reports the homology-based RT-PCR cloning and functional expression of a Gersemia fruticosa analogue of the allene oxide synthase-lipoxygenase (AOS-LOX) fusion protein. The G. fruticosa mRNA codes for a protein with 84% sequence identity to the P. homomalla AOS-LOX. Our data indicate that the AOS-LOX fusion protein pathway is used by another coral and P. homomalla represents no exception.

Source: Biochim Biophys Acta. (2008) vol. 1780, p. 315-321

Cold and water stresses produce changes in endogenous jasmonates in two populations of Pinus pinaster Ait

2007-09-29T19:34:00.002+09:00

There is considerable evidence suggesting that jasmonates (JAs) play a role in plant resistance against abiotic stress. It is well known that in Angiosperms JAs are involved in the defense response, however there is little information about their role in Gymnosperms. Our proposal was to study the involvement of JAs in Pinus pinaster Ait. reaction to cold and water stress, and to compare the response of two populations of different provenances (Gredos and Bajo Tiétar) to these stresses. We detected 12-oxo-phytodienoic acid (OPDA), jasmonic acid (JA), and the hydroxylates 11-hydroxyjasmonate and 12-hydroxyjasmonate in foliage and shoots of P. pinaster plants. The response of the Gredos population to cold stress differed from that of Bajo Tiétar. Gredos plants showed a lower JA-basal level than Bajo Tiétar; under cold stress JA increased twofold at 72 h, while it decreased in Bajo Tiétar plants. The hydroxylates slightly increased in both populations due to cold stress treatment. Under water stress, plants from Gredos showed a remarkable JA-increase; thus the JA-response was much more prominent under water stress than under cold stress. In contrast, no change was found in JA-level in Bajo Tiétar plants under water stress. The level of JA-precursor, OPDA, was very low in control plants from Gredos and Bajo Tiétar. Under water stress OPDA increased only in plants from Bajo Tiétar. Therefore, we inform here of a different JAs-accumulation pattern after the stress treatment in P. pinaster from two provenances, and suggest a possible correlation with adaptations to diverse ecological conditions.

Source: Plant Growth Regulation (2007) vol. 52, p.111-116

The Jasmonate-Induced Expression of the Nicotiana tabacum Leaf Lectin

2007-08-25T21:32:00.002+09:00

Previous experiments with tobacco (Nicotiana tabacum L. cv Samsun NN) plants revealed that jasmonic acid methyl ester (JAME) induces the expression of a cytoplasmic/nuclear lectin in leaf cells and provided the first evidence that jasmonates affect the expression of carbohydrate-binding proteins in plant cells. To corroborate the induced accumulation of relatively large amounts of a cytoplasmic/nuclear lectin, a detailed study was performed on the induction of the lectin in both intact tobacco plants and excised leaves. Experiments with different stress factors demonstrated that the lectin is exclusively induced by exogeneously applied jasmonic acid and JAME, and to a lesser extent by insect herbivory. The lectin concentration depends on leaf age and the position of the tissue in the leaf. JAME acts systemically in intact plants but very locally in excised leaves. Kinetic analyses indicated that the lectin is synthesized within 12 h exposure time to JAME, reaching a maximum after 60 h. After removal of JAME, the lectin progressively disappears from the leaf tissue. The JAME-induced accumulation of an abundant nuclear/cytoplasmic lectin is discussed in view of the possible role of this lectin in the plant.

Source: Plant and cell physiology (2007) vol. 48, p. 1207-1218

Expression of an NADP-malic enzyme gene in rice (Oryza sativa. L) is induced by environmental stresses; over-expression of the gene in ...

2007-07-14T21:26:00.002+09:00

Expression of an NADP-malic enzyme gene in rice (Oryza sativa. L) is induced by environmental stresses; over-expression of the gene in Arabidopsis confers salt and osmotic stress tolerance

NADP-malic enzyme (NADP-ME, EC 1.1.1.40) functions in many different pathways in plants, and has recently been implicated in plant defense such as in responses to wounding and UV-B radiation. In this study, we isolated a complementary DNA (cDNA) clone by using the differential display method and screening of a root cDNA library of rice (Oryza sativa. L) under carbonate (NaHCO3) stress, and identified it as one of the rice NADP-ME genes (we named it NADP-ME2, GenBank accession no. AB053295). The 5' end of NADP-ME2 was obtained by the 5'-RACE method, and the full-length cDNA had a length of 2217 bp encoding 593 amino acids. Expression of NADP-ME2 mRNA in roots was induced by stress from carbonates (NaHCO3 and Na2CO3, NaCl, and environmental pH changes. NADP-ME2 transcripts increased during 72-h exposures to NaHCO3, NaCl, and PEG stresses. Furthermore, NADP-ME activities in leaves and roots of rice seedlings increased by more than 50% in the presence of carbonates (NaHCO3 and Na2CO3), NaCl, and PEG. These results indicate that rice NADP-ME2 responds to salts and osmotic stresses. Transgenic Arabidopsis plants over-expressing NADP-ME2 were obtained through transformation, screening, Northern analysis and in situ NADP-ME activity assay. Transgenic Arabidopsis plants over-expressing NADP-ME2 grew well in 1/2 x MS medium with 100 mM NaCl or 4% mannitol, whereas growth of wild-type (WT) Arabidopsis seedlings was strongly inhibited. In addition, under 125 mM NaCl stress, the root lengths of transgenic lines were about twice as long as those of the WT. These results suggest that NADP-ME2 has a role in enhancing tolerance of plants to salt and osmotic stress.

Source: Plant Mol Biol. (2007) vol. 64, p. 49-58

Enhancing salt tolerance in a crop plant by overexpression of glyoxalase II

2007-05-19T21:24:00.002+09:00

Earlier we have shown the role of glyoxalase overexpression in conferring salinity tolerance in transgenic tobacco. We now demonstrate the feasibility of same in a crop like rice through overproduction of glyoxalase II. The rice glyoxalase II was cloned in pCAMBIA1304 and transformed into rice (Oryza sativa cv PB1) via Agrobacterium. The transgenic plants showed higher constitutive activity of glyoxalase II that increased further upon salt stress, reflecting the upregulation of endogenous glyoxalase II. The transgenic rice showed higher tolerance to toxic concentrations of methylglyoxal (MG) and NaCl. Compared with non-transgenics, transgenic plants at the T1 generation exhibited sustained growth and more favorable ion balance under salt stress conditions.

Source: Transgenic Res. (2008) vol. 17, p. 171-180

Wounding increases salt tolerance in tomato plants: evidence on the participation of calmodulin-like activities in cross-tolerance signalling

2007-03-17T10:46:00.003+09:00

Cross-tolerance is the phenomenon by which a plant resistance to a stress results in resistance to another form of stress. It has previously been shown that salt stress causes the accumulation of proteinase inhibitors and the activation of other wound-related genes in tomato plants (Solanum lycopersicum). However, very little is known about how different stresses interact with one another, and which are the signalling components that interrelate the responses triggered by different stress types. In the present work, it is shown that mechanical wounding increases salt-stress tolerance in tomato plants through a mechanism that involves the signalling peptide systemin and the synthesis of JA. Data are also provided indicating that calmodulin-like activities are necessary for the downstream signalling events that lead to cross-tolerance between wounding and salt stress. Finally, evidence was gathered supporting the hypothesis that LeCDPK1, a Ca2+-dependent protein kinase from tomato previously described in our laboratory, could participate in this cross-tolerance mechanism interrelating the signalling responses to wounding and salt stress.

Source: Journal of Experimental Botany (2006) vo.. 57, p. 2391-2400

Hydrogen peroxide is involved in methyl jasmonate-induced senescence of rice leaves

2007-01-13T10:45:00.003+09:00

The role of H2O2 in the senescence of detached rice leaves induced by methyl jasmonate (MJ) was investigated. MJ treatment resulted in H2O2 production in detached rice leaves, which was prior to the occurrence of leaf senescence. Dimethylthiourea, a chemical trap of H2O2, was observed to be effective in inhibiting MJ-induced senescence and MJ-increased malondialdehyde (MDA) content in detached rice leaves. Diphenyleneiodonium chloride (DPI) and imidazole (IMD), inhibitors of NADPH oxidase, prevented MJ-induced H2O2 production, suggesting that NADPH oxidase is a H2O2-generating enzyme in MJ-treated detached rice leaves. DPI and IMD also inhibited MJ-promoted senescence and MJ-increased MDA content in detached rice leaves. Phosphatidylinositol 3-kinase inhibitors wortmannin (WM) or LY 294002 (LY) inhibited MJ-induced H2O2 production and senescence of detached rice leaves. Exogenous H2O2 reversed the inhibitory effect of WM or LY. In terms of leaf senescence, it was observed that rice seedlings of cultivar Taichung Native 1 (TN1) are jasmonic acid (JA)-sensitive and those of cultivar Tainung 67 (TNG67) are JA-insensitive. On treatment with JA, H2O2 accumulated in the leaves of TN1 seedlings but not in the leaves of TNG67. Evidence was also provided to show that MJ-induced H2O2 production in detached rice leaves is abscisic acid (ABA)-independent. Ethylene action inhibitor, silver thiosulfate, was observed to inhibit MJ- and ABA-induced H2O2 production and senescence of detached rice leaves, suggesting that the action of MJ and ABA is ethylene-dependent.

Source: Physiologia Plantarum (2006) vol. 127, p. 293-303

Duplicate maize 13-lipoxygenase genes are differentially regulated by circadian rhythm, cold stress, wounding, pathogen infection, and hormonal treatm

2006-12-09T00:34:00.003+09:00

Most plant oxylipins, a large class of diverse oxygenated polyunsaturated fatty acids and their derivatives, are produced through the lipoxygenase (LOX) pathway. Recent progress in dicots has highlighted the biological roles of oxylipins in plant defence responses to pathogens and pests. By contrast, the physiological function of LOXs and their metabolites in monocots is poorly understood. Two maize LOXs, ZmLOX10 and ZmLOX11 that share >90% amino acid sequence identity but are localized on different chromosomes, were cloned and characterized. Phylogenetic analysis revealed that ZmLOX10 and ZmLOX11 cluster together with well-characterized plastidic type 2 linoleate 13-LOXs from diverse plant species. Regio-specificity analysis of recombinant ZmLOX10 protein overexpressed in Escherichia coli proved it to be a linoleate 13-LOX with a pH optimum at pH 8.0. Both predicted proteins contain putative transit peptides for chloroplast import. ZmLOX10 was preferentially expressed in leaves and was induced in response to wounding, cold stress, defence-related hormones jasmonic acid (JA), salicylic acid (SA), and abscisic acid (ABA), and inoculation with an avirulent strain of Cochliobolus carbonum. These data suggested a role for this gene in maize adaptation to abiotic stresses and defence responses against pathogens and pests. ZmLOX11 was preferentially expressed in silks and was induced in leaves only by ABA, indicating its possible involvement in responses to osmotic stress. In leaves, mRNA accumulation of ZmLOX10 is strictly regulated by a circadian rhythm, with maximal expression coinciding temporally with the highest photosynthetic activity. This study reveals the evolutionary divergence of physiological roles for relatively recently duplicated genes. Possible physiological functions of these 13-LOXs are suggested.

Source: J Exp Bot. (2006) vol. 57, 3767-3779

Expression of yeast SOD2 in transgenic rice results in increased salt tolerance

2006-11-04T00:33:00.003+09:00

Agricultural productivity is severely affected by soil salinity. One possible mechanism by which plants could survive salt stress is to remove sodium ions from the cytosol via plasma membrane Na+/H+ antiporters. In the present study, we demonstrated that expressing the plasma membrane Na+/H+ antiporter SOD2 from yeast (Schizosaccharomyces pombe) in transgenic rice increased salt tolerance. These transgenic plants accumulated more K+, Ca2+, Mg2+ and less Na+ in their shoots compared with those of non-transformed controls. Moreover, measurements on isolated plasma membrane vesicles derived from the SOD2 transgenic rice plant roots showed that the vesicles had enhanced P-ATPase hydrolytic activity. Furthermore, the transformed rice plants maintained higher levels of photosynthesis and root proton exportation capacity, whereas reduced ROS generation. Physiological analysis suggested that transgenic rice plants might employ multiple mechanisms to improve their salt tolerance under salt stress conditions.

Source: Plant science (2006) vol. 170, p. 216-224

Identification of a peroxisomal acyl-activating enzyme involved in the biosynthesis of jasmonic acid in Arabidopsis

2006-09-23T00:33:00.003+09:00

Jasmonic acid (JA) is a lipid-derived signal that regulates a wide variety of developmental and defense-related processes in higher plants. JA is synthesized from linolenic acid via an enzymatic pathway that initiates in the plastid and terminates in peroxisomes. The C18 JA precursor 12-oxo-phytodienoic acid (OPDA) is converted in the peroxisome to 3-oxo-2-(2'-[Z]-pentenyl)cyclopentane-1-octanoic acid (OPC-8:0), which subsequently undergoes three rounds of beta-oxidation to yield JA. Although most JA biosynthetic enzymes have been identified, several key steps in the pathway remain to be elucidated. To address this knowledge gap, we employed co-expression analysis to identify genes that are coordinately regulated with known JA biosynthetic components in Arabidopsis. Among the candidate genes uncovered by this approach was a 4-coumarate-CoA ligase-like member of the acyl-activating enzyme (AAE) gene family, which we have named OPC-8:0 CoA Ligase1 (OPCL1). In response to wounding, opcl1 null mutants exhibited reduced levels of JA and hyperaccumulation of OPC-8:0. Recombinant OPCL1 was active against both OPDA and OPC-8:0, as well as medium-to-long straight-chain fatty acids. Subcellular localization studies with green fluorescent protein-tagged OPCL1 showed that the protein is targeted to peroxisomes. These findings establish a physiological role for OPCL1 in the activation of JA biosynthetic precursors in leaf peroxisomes, and further indicate that OPC-8:0 is a physiological substrate for the activation step. The results also demonstrate the utility of co-expression analysis for identification of factors that contribute to jasmonate homeostasis.

Source: J Biol Chem. (2006) vol. 281, p. 33511-33520

Lipoxygenases during Brassica napus seed germination

2006-08-05T00:32:00.003+09:00

The peroxidation of polyunsaturated fatty acids is mostly catalyzed by members of the lipoxygenase enzyme family. Lipoxygenase products can be metabolized further in the oxylipin pathway and are known as signalling substances that play a role in plant development as well as in plant responses to wounding and pathogen attack. Apart from accumulating data in model plants like Arabidopsis, information on the relevance of lipid peroxide metabolism in the crop plant oilseed rape is scarce. Thus we aimed to analyze lipoxygenases and oxylipin patterns in seedlings of oilseed rape. RNA isolated from 3 day etiolated seedlings contains mRNAs for at least two different lipoxygenases. These have been cloned as cDNAs and named Bn-Lox-lfl and Bn-Lox-2fl. The protein encoded by Bn-Lox-2fl was identified as a 13-lipoxygenase by expression in Escherichia coli. The Bn-Lox-lfl yielded an inactive protein when expressed in E. coli. Based on Bn-Lox-lfl active site determinants and on sequence homology the Bn-Lox-lfl is most likely a 9-lipoxygenase. Both genes are expressed in light-grown and etiolated cotyledons as well as in leaves. Bn-Lox-2fl protein is more abundant in cotyledons of etiolated seedlings than in cotyledons of green seedlings. Both 13- and 9-lipoxygenase-derived hydroperoxides can be detected during germination. Etiolated seedlings contain more lipoxygenase-derived hydroperoxides in non esterified fatty acids than green seedlings. The 13-lipoxygenase derivatives are 6-8-fold more abundant than the 9-derivatives. Lipoxygenase-derived hydroperoxides in esterified lipids are almost not present during germination. These results suggest that 13-lipoxygenases acting on free fatty acids dominate during B. napus seed germination.

Source: Phytochemistry (2006) vol. 67, p. 2030-2040

Over-expression of chloroplastic lipoxygenase RCI1 causes PR1 transcript accumulation in transiently transformed rice

2006-06-24T00:32:00.003+09:00

The acquired resistance state of rice plants, rendering them resistant to subsequent infections with virulent pathogens, can be triggered by treatments with non-host pathogens, necrotizing pathogens and certain chemicals. The 13-lipoxygenase RCI1 (LOX2:Os:2) is induced specifically following treatment with chemical inducers of resistance. Here, we report that the over-expression of RCI1 in a transient transformation assay leads to PR1 transcript accumulation in rice leaves. In addition, we show that this property is due to the enzymatic activity of the transgenic protein. Furthermore, exogenous application of (13S,92,11E,15Z)-13-hydroxy-9,11,15-octadecatrienoic acid (13-HOT), an oxylipin deriving from the reductase branch of the lipoxygenase pathway, is capable of inducing PR1. These results support a role for RCI1 in the generation of signalling molecules during the establishment of the acquired resistance response in rice.

Source: Physiological and molecular plant pathology (2004) vol. 64, p. 37-43

The outcomes of concentration-specific interactions between salicylate and jasmonate signaling include synergy, antagonism, and oxidative...

2006-04-29T00:31:00.003+09:00

The outcomes of concentration-specific interactions between salicylate and jasmonate signaling include synergy, antagonism, and oxidative stress leading to cell death

Salicylic acid (SA) has been proposed to antagonize jasmonic acid (JA) biosynthesis and signaling. We report, however, that in salicylate hydroxylase-expressing tobacco (Nicotiana tabacum) plants, where SA levels were reduced, JA levels were not elevated during a hypersensitive response elicited by Pseudomonas syringae pv phaseolicola. The effects of cotreatment with various concentrations of SA and JA were assessed in tobacco and Arabidopsis (Arabidopsis thaliana). These suggested that there was a transient synergistic enhancement in the expression of genes associated with either JA (PDF1.2 [defensin] and Thi1.2 [thionin]) or SA (PR1 [PR1a--glucuronidase in tobacco]) signaling when both signals were applied at low (typically 10–100 µM) concentrations. Antagonism was observed at more prolonged treatment times or at higher concentrations. Similar results were also observed when adding the JA precursor, -linolenic acid with SA. Synergic effects on gene expression and plant stress were NPR1- and COI1-dependent, SA- and JA-signaling components, respectively. Electrolyte leakage and Evans blue staining indicated that application of higher concentrations of SA + JA induced plant stress or death and elicited the generation of apoplastic reactive oxygen species. This was indicated by enhancement of hydrogen peroxide-responsive AoPR10--glucuronidase expression, suppression of plant stress/death using catalase, and direct hydrogen peroxide measurements. Our data suggests that the outcomes of JA-SA interactions could be tailored to pathogen/pest attack by the relative concentration of each hormone.

Source: Plant Physiology (2006) vol. 140, p. 249-262

Enhanced drought tolerance of transgenic rice plants expressing a pea manganese superoxide dismutase

2006-03-11T00:30:00.003+09:00

We investigated the role that manganese superoxide dismutase (MnSOD), an important antioxidant enzyme, may play in the drought tolerance of rice. MnSOD from pea (Pisum sativum) under the control of an oxidative stress-inducible SWPA2 promoter was introduced into chloroplasts of rice (Oryza sativa) by Agrobacterium-mediated transformation to develop drought-tolerant rice plants. Functional expression of the pea MnSOD in transgenic rice plants (T1) was revealed under drought stress induced by polyethylene glycol (PEG) 6000. After PEG treatment the transgenic leaf slices showed reduced electrolyte leakage compared to wild type (WT) leaf slices, whether they were exposed to methyl viologen (MV) or not, suggesting that transgenic plants were more resistant to MV- or PEG-induced oxidative stress. Transgenic plants also exhibited less injury, measured by net photosynthetic rate, when treated with PEG. Our data suggest that SOD is a critical component of the ROS scavenging system in plant chloroplasts and that the expression of MnSOD can improve drought tolerance in rice.

Source: J Plant Physiol. (2005) vol. 162, p. 465-472

OsCDPK13, a calcium-dependent protein kinase gene from rice, is induced by cold and gibberellin in rice leaf sheath

2006-01-07T00:30:00.003+09:00

Calcium-dependent protein kinases (CDPKs) play an important role in rice signal transduction, but the precise role of each individual CDPK is still largely unknown. Recently, a full-length cDNA encoding OsCDPK13 from rice seedling was isolated. To characterize the function of OsCDPK13, its responses to various stresses and hormones were analyzed in this study. OsCDPK13 accumulated in 2-week-old leaf sheath and callus, and became phosphorylated in response to cold and gibberellin (GA). OsCDPK13 gene expression and protein accumulation were up-regulated in response to GA3 treatment, but suppressed in response to abscisic acid and brassinolide. Antisense OsCDPK13 transgenic rice lines were shorter than the vector control lines, and the expression of OsCDPK13 was lower in dwarf mutants of rice than in wild type. Furthermore, OsCDPK13 gene expression and protein accumulation were enhanced in response to cold, but suppressed under salt and drought stresses. Sense OsCDPK13 transgenic rice lines had higher recovery rates after cold stress than vector control rice. The expression of OsCDPK13 was stronger in cold-tolerant rice varieties than in cold-sensitive ones. The results suggest that OsCDPK13 might be an important signaling component in the response of rice to GA and cold stress.

Source: Plant Mol Biol. (2004) vol. 55, p. 541-552

Accumulation of SALT protein in rice plants as a response to environmental stresses

2005-08-13T00:25:00.003+09:00

Germination and subsequent hydroponic growth under salt stress (100 mmol/L NaCl) triggered an accumulation of six major stress proteins and resulted in a growth arrest of young seedlings of rice (Oryza sativa L.) cv. Bura Rata. Based on two-dimensional electrophoretic resolution, partial amino acid sequencing and immunodetection techniques, four of the salt stress-induced polypeptides were identified as LEA proteins. Under all experimental conditions wherein seedlings exhibited superior halotolerance, salt stress-induced LEA proteins were expressed at low levels. In contrast, accumulation of LEA proteins was found associated with growth arrest. When returned to non-saline media, seedlings stressed with salt for four days recovered immediately. Longer exposure to 100 mmol/L NaCl, however, progressively delayed recovery and reduced the number of seedlings which could recover from salt stress. Recovery from salt stress was consistently accompanied by degradation of the salt stress-induced LEA proteins. The results of this study show that LEA proteins accumulate during the salinity-triggered growth arrest of young Bura Rata seedlings and are mobilised during the recovery of seedlings from salinity stress.

Source: Journal of Plant Physiology (2003) vol. 160, p. 1165-1174

Development of transgenic rice plants overexpressing the Arabidopsis H+/Ca2+ antiporter CAX1 gene

2005-07-23T00:25:00.003+09:00

The gene of the Arabidopsis thaliana H+/Ca2+ transporter, CAX1 (cation exchanger 1) was introduced into Japonica cultivars of rice (Ilpumbyeo) by Agrobacterium-mediated transformation, and a large number of transgenic plants were produced. The neomycin phosphotransferase II (NPTII) gene was used as a selectable marker. The activity of neomycin phosphotransferase could be successfully detected in transgenic rice callus. The introduction of the CAX1 gene was also proven by PCR using CAX1-specific oligonucleotide primers in regenerated plants. Stable integration and expression of the CAX1 gene in T0 plants and T1 progeny were confirmed by DNA hybridization, Northern blot analysis, and luminescent analysis.

Source: Plant Cell Rep. (2005) vol. 23, p. 678-682

Inheritance and expression of the cry1Abgene in Bt ( Bacillus thuringiensis) transgenic rice

2005-07-02T00:24:00.003+09:00

The inheritance and expression patterns of the cry1Ab gene were studied in the progenies derived from different Bt (Bacillus thuringiensis) transgenic japonica rice lines under field conditions. Both Mendelian and distorted segregation ratios were observed in some selfed and crossed F2 populations. Crosses between japonica intrasubspecies had no significant effect on the segregation ratios of the cry1Ab gene, but crossing between japonica and indica inter-subspecies led to distorted segregation of the cry1Ab gene in the F2 population. Field-release experiments indicated that the cry1Ab gene was stably transmitted in an intact manner via successive sexual generations, and the concentration of the CrylAb protein was kept quantitatively stable up to the R6 generation. The cry1Ab gene, driven by the maize ubiquitin promoter, displayed certain kinds of spatial and temporal expression patterns under field conditions. The content of the CrylAb protein varied in different tissues of the main stems, the primary tillers and the secondary tillers. Higher levels of the CrylAb protein were found in the stems, leaves and leaf sheaths than in the roots, while the lowest level was detected in grains at the maturation stage. The content of the Cry1Ab protein in the leaves peaked at the booting stage and was lowest at the heading stage. Furthermore, the CrylAb content of cry1Ab expression in different tissues of transgenic rice varied individually with temperature.

Source: Theoretical and Applied Genetics (2002) vol. 104, p. 727-734

Scavenging of reactive oxygen species in NaCl-stressed rice (Oryza sativa L.)—differential response in salt-tolerant and sensitive varieties

2005-06-18T00:24:00.003+09:00

Salinity stress affects the metabolism of plant cells leading to severe crop damage and loss of productivity. Oxidative stress is one consequence of salinity that may be responsible for much of the damage. We investigated the immediate responses (enzymatic and non-enzymatic) to salinity-induced oxidative stress in two major rice (Oryza sativa L.) cultivars, salt sensitive Pusa Basmati I (PB) and salt-tolerant Pokkali (PK). Seedlings of both cultivars were subjected to NaCI stress (100-300 mM) for 42 h. Under NaCI stress, the salt-tolerant cv. PK showed higher activity of the ROS scavenging enzyme, catalase (CAT) and enhanced levels of antioxidants like ascorbate (ASC) and glutathione (GSH), than the sensitive cv. PB. Although superoxide dismutase (SOD) activity was lower in cv. PK, it showed lesser extent of membrane damage (lipid peroxidation) and lower levels of H2O2 than cv. PB under stress. The high levels of catalase activity indicate efficient scavenging of H2O2, which is produced more by non-enzymatic means than via SOD in cv. PK. These data indicate that concerted action of both enzymatic and non-enzymatic ROS scavenging machineries is vital to overcome salinity-induced oxidative stress in rice.

Source: Plant science (2003) vol. 165,p. 1411-1418

Agrobacterium-mediated transformation of Indica rice genotypes: an assessment of factors affecting the transformation efficiency

2005-06-04T00:23:00.003+09:00

An efficient Agrobacterium-mediated method for transformation of popular Bangladeshi Indica rice genotypes has been developed. Mature embryo-derived calluses as well as immature embryos were used as the target material. Transgenic plant production frequency was higher using the immature embryos than mature embryo-derived calluses. However, 3-week-old mature embryo-derived calluses served as an excellent starting material. The super-binary vector (pTOK233) was generally more effective than the binary vector (pC1301-Xa21mSS) particularly with recalcitrant Bangladeshi genotypes such as BR22. However, transformation of the Japonica cultivar Taipei-309 was equally effective with either plasmid. Inclusion of acetosyringone (200M) in co-cultivation media proved essential for successful transformation and the optimum co-cultivation period found was to be 3days. A large number of morphologically normal, fertile transgenic plants were obtained which expressed gus as determined by histochemical staining. Integration of the hpt gene into the genome of transgenic plants was confirmed by molecular analysis. Mendelian inheritance of transgenes (hpt and gus gene) was observed in T1 progeny.

Source: Plant Cell, Tissue and Organ Culture (2005) vol. 82, p. 45-55

Stress-inducible synthesis of proline in transgenic rice confers faster growth under stress conditions than that with constitutive synthesis

2005-05-07T00:22:00.004+09:00

Proline accumulation has been shown to correlate with tolerance to drought and salt stresses in plants. Our goal was to compare the growth rate of transgenic rice plants in which the expression of a mothbean Δ1-pyrroline-5-carboxylate synthetase (p5cs) cDNA was driven separately with a constitutive and a stress-inducible promoter. We found that both constitutive expression and stress-inducible expression of the p5cs cDNA in transgenic rice have led to the accumulation of p5cs mRNA and proline. Third-generation (R2) transgenic rice seedlings showed significantly higher tolerance to stress produced by high levels of NaCl or water deficiency as judged by faster growth of shoots and roots in comparison with non-transformed plants. However, stress-inducible expression of the P5CS transgene showed significant advantages over the constitutive expression in increasing the biomass production of transgenic rice grown in soil under stress conditions.

Source: Plant Science (2004) Vol. 166, p. 941-948

Expression of ascorbate peroxidase and glutathione reductase in roots of rice seedlings in response to NaCl and H2O2

2005-04-09T00:22:00.003+09:00

The accumulation of H2O2 by NaCl was observed in the roots of rice seedlings. Treatment with NaCl caused an increase in the activities of ascorbate peroxidase (APX) and glutathione reductase (GR) and the expression of OsAPX and OsGR in rice roots. Exogenously applied H2O2 also enhanced the activities of APX and GR and the expression of OsAPX and OsGR in rice roots. The accumulation of H2O2 in rice roots in response to NaCl was inhibited by the NADPH oxidase inhibitors, diphenyleneiodonium chloride (DPI) and imidazole (IMD). However, DPI, IMD, and dimethylthiourea, a H2O2 trap, did not reduce NaCl-enhanced activities of APX and GR and expression of OsAPX and OsGR. It appears that H2O2 is not involved in the regulation of NaCl-induced APX and GR activities and OsAPX and OsGR expression in rice roots.

Source: Journal of plant physiology (2005) vol. 162, p. 291-299

Over-expression of ζ glutathione S-transferase in transgenic rice enhances germination and growth at low temperature

2005-03-26T00:21:00.003+09:00

To develop a rice cultivar that would be suitable for direct-seedingcultivation in cooler temperate regions, we generated transgenic rice plants inwhich a rice encoding a -class glutathioneS-transferase (GST) under the control of a maize ubiquitinpromoter. GSTs have been suggested to be responsible for tolerance to variousstresses such as cold, salt and drought by detoxification of xenobioticcompounds and reactive oxygen species. A total of 87 R0 transgenicrice plants harboring a chimeric GST gene were generatedusing Agrobacterium mediated transformation. ThreeR2 lines homozygous for the transgene were assayed for GST activityand had higher GST and glutathione peroxidase activities thannon-transformants.Seedlings of the transgenic lines demonstrated greatly enhanced germination andgrowth rates at low temperature grown under submergence. The GST transgeniclines should be useful for breeding rice cultivars suitable for direct-seedingcultivation in cooler temperate regions.

Source: Molecular Breeding (2002) vol. 9, p. 93-101

Optimising the tissue culture conditions for high efficiency transformation of indica rice

2005-02-26T00:20:00.003+09:00

Establishment of high efficiency Agrobacterium-mediated transformation techniques has greatly accelerated the widespread application of transformation in japonica rice. However, transformation in indica rice remains difficult. In this study, we identify two new media for subculture and differentiation, the two major steps in the tissue culture process for transformation. These media were tested using four cultivars representing very different germplasms of indica rice. The results show that the new media significantly improved the growth rate and quality of the calli, and also increased the differentiation rate for all four cultivars tested. Use of these modified media in transformation experiments also greatly improved the transformation efficiency of all four indica cultivars.

Source: Plant Cell Rep. (2005) vol. 23, p. 540-547

Variability in expression of insecticidal Cry1Ab gene in Indica Basmati rice

2005-01-15T00:18:00.004+09:00

The expression of an insecticidal gene cry1Ab, under three different promoters was studied in leaves, stem and panicles to determine organ-specificity in Basmati rice. Enhanced resistance against two Lepidopteran insects, stem borer (Scirpophaga incertulas) and leaf folder (Cnaphalocrocis medinalis) was observed. The result of western hybridization and insect bioassays demonstrated that all these promoters express the cry1Ab gene at similar levels in leaves and panicles. The cry1Ab gene was expressed in stems at 0.05% of the total protein under the control of the PEPC promoter alone or in combination with the pollen-specific promoter. On the other hand it was expressed at 0.15% under the control of the ubiquitin promoter. Southern blot hybridization of these plants indicated integration of the complete plant transcriptional unit at multiple insertion sites. These results demonstrated that a specific promoter could be used to limit the expression of cry1Ab gene in the desired parts of Basmati rice plants.

Source: Euphytica (2002) vol. 128, p. 121-128

Over-expression of the bacterial nhaA gene in rice enhances salt and drought tolerance

2004-12-20T23:57:00.004+09:00

The Escherichia coli nhaA gene encodes a Na+/H+ antiporter, which plays a critical role in ion homeostasis. We transferred a bacterial nhaA gene into rice (Oryza sativa L. ssp. japonica) and detected high expression in the transgenic rice. The germination rate, growth, and average yield per plant of the transgenic lines were better than those of control lines under salt or drought stress. Moreover, the sodium and proline content of the transgenic lines under salt or drought stress was also higher than in control lines, implying that nhaA over-expression enhances osmoregulation by activating the biosynthesis of proline. Tolerance to both salt and drought was compared between transgenic rice over-expressing nhaA and that over-expressing Arabidopsis δ-OAT encoding ornithine-8-aminotransferase. The transgenic plants with nhaA grew better than those with δ-OAT at high salinity, while the opposite was true for drought stress.

Source: Plant science (2005) vol. 168, p. 297-302

Heat-tolerant basmati rice engineered by over-expression of hsp101

2004-09-18T16:37:00.002+09:00

Rice is sensitive to high-temperature stress at almost all the stages of its growth and development. Considering the crucial role of heat shock protein 101 (Hsp 101) in imparting thermotolerance to cells, we introduced Arabidopsis thaliana hsp101 (Athsp101) cDNA into the Pusa basmati 1 cultivar of rice (Oryza sativa L.) by Agrobacterium-mediated transformation. Stable integration and expression of the transgene into the rice genome was demonstrated by Southern, northern and western blot analyses. There appeared no adverse effect of over-expression of the transgene on overall growth and development of transformants. The genetic analysis of tested T1 lines showed that the transgene segregated in a Mendelian fashion. We compared the survival of T2 transgenic lines after exposure to different levels of high-temperature stress with the untransformed control plants. The transgenic rice lines showed significantly better growth performance in the recovery phase following the stress. This thermotolerance advantage appeared to be solely due to over-expression of Hsp101 as neither the expression of low-molecular-weight heat shock proteins (HSPs) nor of other members of Clp family proteins was altered in the transgenic rice. The production of high temperature tolerant transgenic rice cultivars would provide a stability advantage under supra-optimal temperature regime thereby improving its overall performance.

Source: Plant Mol Biol. (2003) vol 51, p. 677-686

Transgenics of an elite indica rice variety Pusa Basmati 1 harbouring the codA gene are highly tolerant to salt stress

2004-08-28T16:33:00.002+09:00

Transgenic lines of indica rice were generated by Agrobacterium-mediated transformation with the choline oxidase ( codA) gene from Arthrobacter globiformis. Choline oxidase catalyses conversion of choline to glycine betaine. Glycine betaine is known to provide tolerance against a variety of stresses. Molecular analyses of seven independent transgenic lines as performed by Southern, Northern and Western hybridization revealed integration and expression of the transgene as well as inheritance in the progeny plants. A good correlation was observed between levels of mRNA and protein accumulation, and a significant amount of choline oxidase product, i.e. glycine betaine, accumulated in R0 as well as R1 plants. Mendelian as well as non-Mendelian segregation patterns were obtained in the progeny plants. Challenge studies performed with R1 plants by exposure to salt stress (0.15 M NaCl) for 1 week, followed by a recovery period, revealed that in some cases more than 50% of the transgenic plants could survive salt stress and set seed whereas wild-type plants failed to recover.

Source: Theor Appl Genet. (2002) vol 106, p. 51-57

Highly efficient production and characterization of T-DNA plants for rice ( Oryza sativa L.) functional genomics

2004-07-11T15:28:00.002+09:00

We investigated the potential of an improved Agrobacterium tumefaciens-mediated transformation procedure of japonica rice ( Oryza sativa L.) for generating large numbers of T-DNA plants that are required for functional analysis of this model genome. Using a T-DNA construct bearing the hygromycin resistance ( hpt), green fluorescent protein ( gfp) and beta-glucuronidase ( gusA) genes, each individually driven by a CaMV 35S promoter, we established a highly efficient seed-embryo callus transformation procedure that results both in a high frequency (75-95%) of co-cultured calli yielding resistant cell lines and the generation of multiple (10 to more than 20) resistant cell lines per co-cultured callus. Efficiencies ranged from four to ten independent transformants per co-cultivated callus in various japonica cultivars. We further analysed the T-DNA integration patterns within a population of more than 200 transgenic plants. In the three cultivars studied, 30-40% of the T(0) plants were found to have integrated a single T-DNA copy. Analyses of segregation for hygromycin resistance in T(1) progenies showed that 30-50% of the lines harbouring multiple T-DNA insertions exhibited hpt gene silencing, whereas only 10% of lines harbouring a single T-DNA insertion was prone to silencing. Most of the lines silenced for hpt also exhibited apparent silencing of the gus and gfp genes borne by the T-DNA. The genomic regions flanking the left border of T-DNA insertion points were recovered in 477 plants and sequenced. Adapter-ligation Polymerase chain reaction analysis proved to be an efficient and reliable method to identify these sequences. By homology search, 77 T-DNA insertion sites were localized on BAC/PAC rice Nipponbare sequences. The influence of the organization of T-DNA integration on subsequent identification of T-DNA insertion sites and gene expression detection systems is discussed.

Source: Theor Appl Genet. (2003) vol 106, p. 1396-1408

Highly efficient production and characterization of T-DNA plants for rice ( Oryza sativa L.) functional genomics

2004-07-10T23:41:00.003+09:00

Trehalose accumulation in rice plants confers high tolerance levels to different abiotic stresses

2004-05-29T23:39:00.004+09:00

Trehalose is a nonreducing disaccharide of glucose that functions as a compatible solute in the stabilization of biological structures under abiotic stress in bacteria, fungi, and invertebrates. With the notable exception of the desiccation-tolerant “resurrection plants,” trehalose is not thought to accumulate to detectable levels in most plants. We report here the regulated overexpression of Escherichia coli trehalose biosynthetic genes (otsA and otsB) as a fusion gene for manipulating abiotic stress tolerance in rice. The fusion gene has the advantages of necessitating only a single transformation event and a higher net catalytic efficiency for trehalose formation. The expression of the transgene was under the control of either tissue-specific or stress-dependent promoters. Compared with nontransgenic rice, several independent transgenic lines exhibited sustained plant growth, less photo-oxidative damage, and more favorable mineral balance under salt, drought, and low-temperature stress conditions. Depending on growth conditions, the transgenic rice plants accumulate trehalose at levels 3–10 times that of the nontransgenic controls. The observation that peak trehalose levels remain well below 1 mg/g fresh weight indicates that the primary effect of trehalose is not as a compatible solute. Rather, increased trehalose accumulation correlates with higher soluble carbohydrate levels and an elevated capacity for photosynthesis under both stress and nonstress conditions, consistent with a suggested role in modulating sugar sensing and carbohydrate metabolism. These findings demonstrate the feasibility of engineering rice for increased tolerance of abiotic stress and enhanced productivity through tissue-specific or stress-dependent overproduction of trehalose.

Source: Proc Natl Acad Sci U S A. (2002) vol 99, p. 15898–15903

Research

2004-04-24T21:17:00.004+09:00

Doctoral Degree (2004 - Now):
* Part I:
1. Production of transgenic rice overexpressing AOS-LOX fusion gene (T0, T1, T2)

2. Genetic Analysis of transgenic rice
- Analysis of DNA (by PCR and/or Southern), RNA (by Northern), Protein (Western)
- Selection of homozygous lines of transgenic rice

3. Biochemical Analysis of transgenic rice
- Lipid analysis
- JA analysis
- Dihydro JA analysis

4. Stress response of overexpressed transgenic rice
- Ion leakage experiments in cold, salt stress
- Wound and HR (local, systemic) response
- Blast fungi resistance

5. Metabolomics of overexpressed transgenic rice
- Lipid peroxidation experiments
- Product analysis by HPLC, GC, GC-MS
- Metabolic changes by wound and other stresses

* Part II:
1. Characterization of Fusion Genes (AOS-LOX, AOS-AOC and AOC-AOS)

Master Degree (2000 - 2002):

1. Production of transgenic rice overexpressing allene oxide synthase gene

2. Kinetic properties of V311M mutant Bacillus subtilis protoporphyrinogene oxidase
- Protein expression
- Protein purification
- Enzyme kinetics

Publications

2004-04-23T21:29:00.008+09:00

Updated on December 22, 2009

Thavrak Huon, Sungkuk Jang, Kyoungwon Cho, Randeep Rakwal, Je Chang Woo, Ilchul Kim, Seung Wook Chi, Oksoo Han (2009) A Substrate Serves as a Hydrogen Atom Donor in the Enzyme-Initiated Catalytic Mechanism of Dual Positional Specific Maize Lipoxygenase-1. Bulletin of the Korean Chemical Society. 30: 719-723 [Abstract] [PDF Full Text]

Sungkuk Jang, Thavrak Huon, Keumhwa Kim, Eunji Um, and Oksoo Han (2007) Regiochemical and stereochemical evidence for enzyme-initiated catalysis in dual positional specific maize lipoxygenase-1. Organic letters 16: 3113-3116 [Abstract] [PDF Full Text]

Kyoungwon Cho, Sungkuk Jang, Thavrak Huon, Sangwook Park and Oksoo Han (2007) Biochemical Characterization of the Dual Positional Specific Maize Lipoxygenase and the Dependence of Lagging and Initial Burst Phenomenon on pH, Substrate, and Detergent during Pre-steady State Kinetics. Journal of Biochemistry and Molecular Biology 40: 100-106 [Abstract] [PDF Full Text]

Eunjoo Jeong, Thavrak Huon, Yongin Kuk, Eun-Seon Kim, Hema Kumar Chandru, Myunggi Baik, Kyoungwhan Back, Ja-Ock Guh, and Oksoo Han (2003) A point mutation of valine-311 to methionine in Bacillus subtilis protoporphyrinogen oxidase does not greatly increase resistance to the diphenyl ether herbicide oxyfluorfen. Bioorganic Chemistry 31: 389-397 [Abstract] [PDF Full Text]

Thavrak Huon (2002) Kinetic properties of V311M mutant Bacillus subtilis protoporphyrinogen oxidase. Thesis of Master degree, Department of Genetic Engineering, Chonnam National University. Gwangju, Republic of Korea

Thavrak Huon (1997) The effect of organic and inorganic fertilization on the growth of Tilapia fingerling. Thesis of Bachelor degree, Faculty of Fisheries, Royal University of Agriculture. Phnom Penh, Cambodia

Photo gallery

2004-04-22T21:45:00.002+09:00

More Photos in 2009

More Photos in 2008

More Photos in 2007

More Photos in 2006

More Photos in 2005

More Photos in 2004

More Photos in 2003

More Photos in 2002

More Photos in 2001

More Photos in 2000

Contact me

2004-04-21T00:25:00.004+09:00

My E-mail: huonth@hotmail.com

My address:

About Me

2004-04-20T01:41:00.005+09:00

Family Name: Huon

First Name: Thavrak

Nationality: Cambodian

Tel: (82) 010-2943-2486

E-mail: huonth@hotmail.com

Homepage: thavrakhuon.blogspot.com

Education Qualifications:

. Doctoral Candidate, Department of Biotechnology (Biochemistry Lab), Chonnam National University (Republic of Korea), March, 2004 - Present

. MSc, Department of Genetic Engineering (Biochemistry Lab), Chonnam National University (Republic of Korea), 2002

. BSc, Faculty of Fisheries, Royal University of Agriculture (Cambodia), 1998