Increased Lysine Content in Crop Plants,Amino Acid,Mutants,Tobacco,Amino Ethyl Cystein,Biosynthetic Pathway,Threonine,Methionine

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Metabolic Engineering and Metabolomics Metabolic Engineering and Metabolomics Introduction Cloning and Expression of Heterologous Genes Completion of Partial Pathways giving new Products Transfer of an Entire Biosynthetic Pathway Creating new Products and new Reactants Altering Nutrient uptake and Metabolite Transfer of Promising Natural Motifs Redirecting Metabolite Flow Desensitizing Feedback Inhibition Increased Threonine in Bacteria Increased Lysine Content in Crop Plants Elevation of the Activity of rate Limiting Enzymes Alteration in Protein Processing Pathway Reduction of Competition for Limited Resources Modification of Metabolic Regulation Molecular Breeding of Biosynthetic Pathways

Home >> Industrial and Microbial Biotechnology >>Metabolic Engineering and Metabolomics >> Increased Lysine Content in Crop Plants

Your Ad Here	Increased lysine content in crop plants Major crops used for human food and animal feed are poor in amino acid content, the lysine being the most limiting amino acid. Corn being poor in lysine, when used as animal feel, is often supplemented with soybean meal. Additional quantity of 200,000 tones of lysine is produced annually (world over) through fermentation to meet the demand of lysine. Therefore, efforts have been made in the past to increase lysine content in food and fodder crops, but conventional breeding was not successful.

Increased Lysine Content in Crop Plants	Your Ad Here
In tobacco, mutants over-producing lysine were obtained through selection for resistance against the lysine analog S (2-amino-ethyl) cysteine (AEC), but increased lysine content was observed only in the leaves and not in the seed. Using biotechnology, successful attempts to increase lysine content were made through deregulation of biosynthetic pathway, which utilizes aspartate and produces lysine, along with threonine, methionine and isoleucine. Lysine synthesis is regulated through feedback inhibition of the following enzymes: aspartokinase (AK) catalyzes the phosphorylation reaction converting aspartate to β aspartyl phosphate and (ii) di-hydro-dipicolinic acid synthase (DHDPS) catalyzes the condensation of aspartyl β-semialdehyde leading to lysine synthesis.
Your Ad Here	Lysine overproduction was achieved by manipulating this pathway with following results in which increased lysine content was observed only in leaves but not in seeds: (i) The enzyme DHDPS from E. coli is 20- fold less sensitive to feedback inhibition than DHDPS of higher plants. Therefore, E. coli gene dapA encoding DHDPS was fused with CaMV 35S promoter and a sequence for chloroplast transit sequence. The gene construct was used for producing transgenic plants in tobacco leading to substantial increase in the lysine content of leaves, but not that of seeds. (ii) A mutant allele of E. coli lysC gene encoding lysine insensitive AK enzyme was also similarly fused with CaMV35S promoter. This was introduced in transgenic plants already carrying E. coli dapA gene. These transgenic tobacco plants carrying both dapA and mutant lysC from E. coli accumulated more lysine than those carrying only dapA but this increase was also observed only in leaves and not in seeds.

(iii) Seed specific promoters were also used with both dapA and the mutant lysC but no increase in lysine in the seeds was observed, which was attributed to increased lysine breakdown (catabolism) in the seed, so that not only more lysine need to be produced or translocated to seeds, but also its breakdown needs to be checked

Following genes were also used for producing high lysine transgenic plants in canola and soybean (i) dapA gene from Corynebacterium encoding lysine insensitive DHDPS and (ii) a mutant lysC-M4 gene encoding lysine insensitive AK. Both these genes were individually fused to cts (from small subunt of ribulose 1,5 bisphosphate carboxylase from soybean) and to a seed specific expression cassette composed of a promoter and transcription terminator from the gene encoding the ß subunit of phaseolin, a seed storage protein from Phaseolus. The transgenic canola plants produced using these gene constructs in some cases, had as much as 100 fold increase in free lysine level (12% of all the amino acids) in seeds. Accumulation of lysine in canola seeds was controlled not only by the rate of its synthesis, but also by the rate of its breakdown. Atleast copies of dapA gene were found to be necessary to achieve excess free lysine.

Transgenic soybean for higher lysine (increase ranged from ten fold to several hundred fold) were also produced using the same two genes, that were used for canola by the same group of workers. Increase in lysine codons in corn seeds was also successfully achieved. In future this approach will also be used for accumulation of other amino acids (e.g. threonine and methionine) derived from aspartate.

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