The responses of L-gulonolactone oxidase and HKT2;1 genes in Aeluropus littoralis' shoots under high concentration of sodium chloride

ABSTRACT: Salinity is one of the most important abiotic stresses that limit crop growth and production. Salt stress influences plants in two ways: by affecting ion toxicity and increasing osmotic stress. Ion homeostasis, the excretion of Na + and using antioxidant systems are the major strategies of salt tolerance in plants. Na + and K + transporters with enzymes that are involved in detoxification of reactive oxygen species play key roles in salt tolerance in plants. The aim of this study was to investigate the responses of high affinity K + transporter2;1 gene (HKT2;1) which is involved in regulation of ion homeostasis and L-gulonolactone oxidase (GLOase) which is involved in the ascorbic acid biosynthesis pathway, under different concentrations of NaCl over different time points in Aeluropus littoralis shoots. Results from Real Time PCR data showed that expressions of both genes were influenced by external and internal concentrations of Na + and the internal K + content. AlHKT2;1 was significantly upregulated by increasing Na + concentration at all time points. Furthermore, its highest expression level in shoots occurred after 6 days in 300mM NaCl in shoots which was 25folds more than untreated shoots. AlGLOase expression levels increased 54 h after initiation of salt stress. These results indicate that AlHKT2;1 and AlGLOase respond to different salinity conditions and probably are part of the mechanisms involved in tolerance to high salt concentrations in A. littoralis

The effects of low-molecular-weight and high-molecular-weight glutenin subunits on physical dough properties of bread wheat / by Farajollah Shahriani-Ahmadi.

Bibliography: leaves 223-249.vi, 249, [9] leaves of plates : ill. ; 30 cm.Demonstrates association between Glu-3 alleles encoding low molecular weight glutenins and the dough properties of wheat flours, as well as the interaction between Glu-3 and Glu-1 (high moilecular weight glutenin) in determing dough properties.Thesis (Ph.D.)--University of Adelaide, Dept. of Plant Science, 199

Transient Expression of cor Gene in Papaver somniferum

Introduction: Papaver somniferum is the commercial source of morphine and codeine. The isolation of effective genes involved in the morphine biosynthesis of P. somniferum is very important in the production of specific metabolites achieved using metabolic engineering techniques. In this pathway, the key enzyme COR is involved in the conversion of codeinone to codeine and morphinone to morphine. Methods: the gene encoding of this enzyme was isolated using primers designed on the base of gene sequence available on (NCBI) for P. somniferum. This gene correct size around (960 bp) was first subcloned into pTZ57RIT vector then cloned into expression vectors (pBI121) between BamHI and SacI sites to allow the expression of cor gene driven by the cauliflower mosaic virus 35S promoter. The result was confirmed through different molecular methods e.g. PCR and enzyme digestion by BamHI and SacI. The recombinant plasmid was transformed into the E. coli strain DH5α using a freeze-thaw method. Having selected positive colones on selection medium, plasmid was extracted by miniprep method and recombinant plasmids were selected based on PCR and digestion. The construct was then mobilized in Agrobacterium tumefaciens C58/pGV3850 (KmR RifR). After gene transformation to P. somniferum plants, the agroinfiltration method was also used for transient expression of COR enzyme. Results: evaluation results showed that morphine and codeine were detectable in the leaves of transgenic plants containing cor transgene and there was significant difference in the final production. After completing this experiment for three times, results showed that in 11 sets from 15 sets of leaves experiment tested, main alkaloids (codeine, morphine, papaverin, noscapine and thebaine) were detectable. Conclusion: Whereas no signal was detected in non-infiltrated control leaves or in leaves infiltrated with non-recombinant bacteria for morphine and codeine, others such as thebaine and papaverine were detectable

Genetically engineered hairy root cultures of Hyoscyamus senecionis and H. muticus: ploidy as a promising parameter in the metabolic engineering of tropane alkaloids

Key Message: Tetraploidy improves overexpression of h6h and scopolamine production of H. muticus, while in H. senecionis, pmt overexpression and elicitation can be used as effective methods for increasing tropane alkaloids. Abstract: The effects of metabolic engineering in a polyploid context were studied by overexpression of h6h in the tetraploid hairy root cultures of H. muticus. Flow cytometry analysis indicated genetic stability in the majority of the clones, while only a few clones showed genetic instability. Among all the diploid and tetraploid clones, the highest level of h6h transgene expression and scopolamine accumulation was interestingly observed in the tetraploid clones of H. muticus. Therefore, metabolic engineering of the tropane biosynthetic pathway in polyploids is suggested as a potential system for increasing the production of tropane alkaloids. Transgenic hairy root cultures of Hyoscyamus senecionis were also established. While overexpression of pmt in H. senecionis was correlated with a sharp increase in hyoscyamine production, the h6h-overexpressing clones were not able to accumulate higher levels of scopolamine than the leaves of intact plants. Applying methyl jasmonate was followed by a sharp increase in the expression of pmt and a drop in the expression of tropinone reductase II (trII) which consequently resulted in the higher biosynthesis of hyoscyamine and total alkaloids in H. senecionis.Peer reviewed: YesNRC publication: Ye

Genetically engineered hairy root cultures of Hyoscyamus senecionis and H. muticus: ploidy as a promising parameter in the metabolic engineering of tropane alkaloids

Key Message: Tetraploidy improves overexpression of h6h and scopolamine production of H. muticus, while in H. senecionis, pmt overexpression and elicitation can be used as effective methods for increasing tropane alkaloids. Abstract: The effects of metabolic engineering in a polyploid context were studied by overexpression of h6h in the tetraploid hairy root cultures of H. muticus. Flow cytometry analysis indicated genetic stability in the majority of the clones, while only a few clones showed genetic instability. Among all the diploid and tetraploid clones, the highest level of h6h transgene expression and scopolamine accumulation was interestingly observed in the tetraploid clones of H. muticus. Therefore, metabolic engineering of the tropane biosynthetic pathway in polyploids is suggested as a potential system for increasing the production of tropane alkaloids. Transgenic hairy root cultures of Hyoscyamus senecionis were also established. While overexpression of pmt in H. senecionis was correlated with a sharp increase in hyoscyamine production, the h6h-overexpressing clones were not able to accumulate higher levels of scopolamine than the leaves of intact plants. Applying methyl jasmonate was followed by a sharp increase in the expression of pmt and a drop in the expression of tropinone reductase II (trII) which consequently resulted in the higher biosynthesis of hyoscyamine and total alkaloids in H. senecionis.Peer reviewed: YesNRC publication: Ye

Production of tropane alkaloids in diploid and tetraploid plants and in vitro hairy root cultures of Egyptian henbane (Hyoscyamus muticus L.)

In this study, the effects of ploidy level and culture medium were studied on the production of tropane alkaloids. We have successfully produced stable tetraploid hairy root lines of Hyoscyamus muticus and their ploidy stability was confirmed 30 months after transformation. Tetraploidy affected the growth rate and alkaloid accumulation in plants and transformed root cultures of Egyptian henbane. Although tetraploid plants could produce 200% higher scopolamine than their diploid counterparts, this result was not observed for corresponding induced hairy root cultures. Culture conditions did not only play an important role for biomass production, but also significantly affected tropane alkaloid accumulation in hairy root cultures. In spite of its lower biomass production, tetraploid clone could produce more scopolamine than the diploid counterpart under similar growth conditions. The highest yields of scopolamine (13.87 mg l−1) and hyoscyamine (107.7 mg 1−1) were obtained when diploid clones were grown on medium consisting of either Murashige and Skoog with 60 g/l sucrose or Gamborg’s B5 with 40 g/l sucrose, respectively. Although the hyoscyamine is the main alkaloid in the H. muticus plants, manipulation of ploidy level and culture conditions successfully changed the scopolamine/hyoscyamine ratio towards scopolamine. The fact that hyoscyamine is converted to scopolamine is very important due to the higher market value of scopolamine