Linoleic Acid Metabolism Key Genes Functional Studies Of BnaATs/BnaOPRs In Brassica Napus L.

The improvement of fatty acid composition is one of the research hotspots in rapeseed quality breeding.Molecular breeding can speed up the breeding process,so it is favored by breeders.The function of key genes has always been the focus of molecular breeding.At present,the related gene studies mainly focus on the major effect gene FADs,but there are few reports about the minor effect gene.In this study,BnaATs and BnaOPRs,key genes related to fatty acid metabolism,were obtained based on mi RNA sequencing,and in-depth studies were carried out.The results are as follows:1.Agrobacterium-mediated over expression and silenced expression vectors of 2copies of BnaATs gene and 4 copies of BnaOPRs gene were transformed into Arabidopsis thaliana,respectively.Linoleic acid content of each transgenic plant:（1）In T₁ generation,the linoleic acid content of overexpressed BnaAT1,BnaAT2,BnaOPR1,BnaOPR2,BnaOPR3 and BnaOPR4 plants increased by 14.11%,15.53%,14.17%,10.02%,14.50%and 11.56%,respectively.The linoleic acid content of silenced plants BnaAT1,BnaAT2,BnaOPR1,BnaOPR2,BnaOPR3 and BnaOPR4decreased by 7.21%,5.67%,10.86%,5.20%,1.58%and 6.26%,respectively.The linoleic acid content of overexpressed BnaAT1,BnaAT2,BnaOPR1,BnaOPR2,BnaOPR3 and BnaOPR4 plants increased by 6.93%,9.73%,9.70%,9.35%,4.47%and 5.22%,respectively.Linoleic acid content of silenced BnaAT1,BnaAT2,BnaOPR1,BnaOPR2,BnaOPR3 and BnaOPR4 reduced by 2.34%,2.55%,0.44%,0.81%,1.14%and 1.01%,respectively.Among them,two copies of BnaAT1 and BnaOPR1 genes significantly improved the linoleic acid content compared with other copies.The obtained T₂ overexpressed A.thaliana material significantly increased the linoleic acid content（6.93%-9.70%）,and the gene silencing significantly reduced the linoleic acid content（about 2.3%）.In addition,the fatty acid components of transgenic plants influenced each other,among which palmitic acid was significantly positively correlated with linoleic acid,oleic acid was significantly negatively correlated with linoleic acid,and oleic acid was significantly negatively correlated with linolenic acid,with correlation coefficients of about 0.9.2.BnaATs and BnaOPRs genes were expressed in lateral branches and leaves,buds,flowers,scones,roots and stems of transgenic A.thaliana,but the highest expression levels were noted in flowers and scones.Abiotic stress studies:（1）after IAA treatment,the expression levels of BnaAT1 and BnaOPR1 were significantly upregulated by 3.6 times and 1.46 times;（2）After NaCl treatment,the expression levels of BnaAT1 and BnaOPR2 were significantly increased by 3.6 and 2.6 times;（3）After GA₃ treatment,the expression levels of BnaAT2 and BnaOPR2 were significantly increased by 33.4 times and 5.3 times;（4）PEG 4000 significantly up-regulated the expression levels of BnaAT2 and BnaOPR4 by 11.4 and 9.15 times.It’s concluded that BnaATs and BnaOPRs genes produced stress response to abiotic stress.3.Subcellular localization of BnaAT1 and BnaOPR1 genes in tobacco showed that BnaAT1 was expressed in the nucleus and cell wall,and BnaOPR1 was expressed in plastids.The over expression vectors of BnaAT1 and BnaOPR1 genes were constructed and successfully transferred into Zhongshuang 11.The real-time PCR（RT-q PCR）results showed that the relative expression levels of BnaAT1 and BnaOPR1 genes were 2.3 and 2.2 times higher than those of untransformed plants,respectively.