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Genetic modification of soybean (Glycine max (L.) Merrill) with delta-6 desaturase gene for gamma linolenic acid production

Sakthivelu, G. (2009) Genetic modification of soybean (Glycine max (L.) Merrill) with delta-6 desaturase gene for gamma linolenic acid production. PhD thesis, University of Mysore.

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Abstract

Engineering of fatty acid pathway in higher plants for enhancing their polyunsaturated fatty acid (PUFA) content is gaining importance in recent years. Gamma linolenic acid (GLA), a long chain omega-6 polyunsaturated fatty acid has been reported to be efficacious in treating nastalgia, atopic eczema, cancer and diabetic neuropathy and the sources are very meager. In the present investigation, studies were undertaken to establish a genetic transformation protocol for soybean and to express the delta-6 desaturase gene in soybean for the production of gamma linolenic acid. In vitro regeneration of soybean was best achieved with cotyledonary nodal explants which gave a maximum of 8 shoots/ explant, under the influence of TDZ. Also addition of triacontanol along with BAP during multiplication and rooting phase increased the number of shoots to 14 per explant, indicating the synergistic effect between them. Embryonic axis served as a better explant for Agrobacterium mediated genetic transformation of soybean resulting in a transformation efficiency of 7.4%, compared to cotyledonary nodes. The full length gene sequence of delta-6 desaturase was isolated from Spirulina platensis (strain Sp-6) and cloned into the binary vector pCAMBIA1305.2 to generate the recombinant plasmid pCAMBIAD6d, under the control of CaMV35S constitutive promoter and 35S poly A terminator. The isolated delta-6 desaturase gene sequence exhibited 98% similarity with the target protein and showed three histidine rich motifs, which may act as an active site for the enzyme activity. Agrobacterium mediated genetic transformation of soybean cv. JS 335 with the recombinant plasmid (pCAMBIAD6d) resulted in the formation of GLA (3.8%) and octadecatetraenoic acid (6.2%) in contrast to their absence in control plants. The results from the present study suggests that delta-6 desaturase gene from Spirulina platensis is quite potential for producing GLA in oilseed crops. Further selection and manipulation of these delta-6 desaturase expressing soybean lines for accumulation of GLA in seeds to a greater extent, may make them nutritionally a more potential oilseed crop.

Item Type: Thesis (PhD)
Uncontrolled Keywords: Fatty acid engineering; Polyunsaturated fatty acid (PUFA); Gamma linolenic acid (GLA); Spirulina platensis; Agrobacterium
Subjects: 500 Natural Sciences and Mathematics > 10 Plants
500 Natural Sciences and Mathematics > 04 Chemistry and Allied Sciences > 17 Fatty Acid Chemistry
600 Technology > 02 Engineering & allied operations
Divisions: Plant Cell Biotechnology
Depositing User: Food Sci. & Technol. Information Services
Date Deposited: 06 Apr 2010 04:58
Last Modified: 06 Apr 2010 04:58
URI: http://ir.cftri.com/id/eprint/9413

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