关键词: 大豆, 种质资源, 脂肪氧化酶, RNA干扰, 重组PCR

Abstract:

【Objective】 The objective of this study was try to find a new avenue of breeding technique in soybean to improve the quality of soybeans and create new superordinary germplasm resource of soybeans by RNAi, which prohibits the gene expression of lipoxygenase in soybean seed. 【Method】 The consensus sequence of lipoxygenase gene (Lx1, Lx2, Lx3) in soybean seeds was cloned by RT-PCR, the length of the sequence was 357 bp. An RNAi expression vector of soybean lipoxygenase gene was reconstructed by recombinant PCR, and transformed the vector in soybean (Jinong 18) by pollen tube pathway. 【Result】 Twelve transgenic strains were obtained and 10 of which demonstrated significant differences. SDS-PAGE and activity of lipoxygenase demonstrated that the lipoxygenase activity of transgenic soybeans decreased significantly, the degree of decreasing was 64.2% on average. The near-infrared analyses demonstrated that the fat content of the transgenic soybeans increased by 0.8%-1.5% and 1.2%-3.0% the total protein decreased by 1.2%-3.0% compared with the control group. The results of RT-PCR demonstrated that the mRNA of lipoxygenase in transgenic soybeans was significantly inhibited. In addition, the activity of lipoxygenase decreased significantly in the progenies of transgenic soybean seeds and the seeds had more fat. 【Conclusion】 The RNAi technology can effectively inhibit the expression of lipoxygenase gene in soybean seed, decrease the activity of lipoxygenase, increase the amount of oil content in its products, improve the quality of soybeans, and create new superordinary germplasm resources of soybean.

Key words: soybean, germplasm resource, lipoxygenase, RNAi, recombinant PCR