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  • Changes in oleic Acid content of transgenic soybeans by antisense RNA mediated posttranscriptional gene silencing.

Changes in oleic Acid content of transgenic soybeans by antisense RNA mediated posttranscriptional gene silencing.

International journal of genomics (2014-09-10)
Ling Zhang, Xiang-Dong Yang, Yuan-Yu Zhang, Jing Yang, Guang-Xun Qi, Dong-Quan Guo, Guo-Jie Xing, Yao Yao, Wen-Jing Xu, Hai-Yun Li, Qi-Yun Li, Ying-Shan Dong
ABSTRACT

The Delta-12 oleate desaturase gene (FAD2-1), which converts oleic acid into linoleic acid, is the key enzyme determining the fatty acid composition of seed oil. In this study, we inhibited the expression of endogenous Delta-12 oleate desaturase GmFad2-1b gene by using antisense RNA in soybean Williams 82. By employing the soybean cotyledonary-node method, a part of the cDNA of soybean GmFad2-1b 801 bp was cloned for the construction of a pCAMBIA3300 vector under the soybean seed promoter BCSP. Leaf painting, LibertyLink strip, PCR, Southern blot, qRT-PCR, and fatty acid analysis were used to detect the insertion and expression of GmFad2-1b in the transgenic soybean lines. The results indicate that the metabolically engineered plants exhibited a significant increase in oleic acid (up to 51.71%) and a reduction in palmitic acid (to <3%) in their seed oil content. No structural differences were observed between the fatty acids of the transgenic and the nontransgenic oil extracts.

MATERIALS
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Roche
DIG-High Prime DNA Labeling and Detection Starter Kit II, sufficient for 12 labeling reactions (10 ng to 3 μg per assay), sufficient for 24 blots (blots of 100 cm2)