山葡萄4CL基因克隆与表达分析  

Expression of the 4CL gene isolated from Vitis amurensis in E.coli

作　　者：陈蒙 刘海峰[1] CHEN Meng;LIU Haifeng(College of Agriculture,Yanbian University,Yanji 133002,China)

机构地区：[1]延边大学农学院

出　　处：《扬州大学学报（农业与生命科学版）》2019年第5期20-25,共6页Journal of Yangzhou University：Agricultural and Life Science Edition

基　　金：国家自然科学基金资助项目(31260067);吉林省教育厅“十三五”科学技术研究规划项目[吉教科合字(2016)第255号]

摘　　要：山葡萄(Vitis amurensis)果皮颜色是影响果实品质的重要因素之一。果皮颜色主要由花色苷决定, 4-香豆酸辅酶A连接酶(4CL)是花色苷合成关键酶。通过RT-PCR技术获得山葡萄4CL基因的全长cDNA为1 772 bp,开放阅读框(ORF)为1 635 bp,编码544个氨基酸多肽,分子量为59.63 ku,等电点为9.02。用RT-qPCR技术,分析4CL在山葡萄果实着色8个发育期的表达差异, 4CL基因表达呈"低-高-低"的变化趋势,果皮全着色(100%)期达峰值,完熟期呈下降趋势。构建原核表达重组质粒pET28a-4CL,并在大肠埃希菌(E.coli) BL21表达4CL,与预期大小一致。The pericarp color of Vitis amurensis is one of the most important factors affecting the quality of the fruit. The pericarp color is maily determined by anthocyanins, and 4-coumarate: coenzyme A ligase(4 CL) is the key enzyme for anthocyanin synthesis. In order to reveal the role of 4 CL in the process of pericarp coloration of V.amurensis, the 4CL gene of V.amurensis in this study was obtained by using RT-PCR, which was 1 772 bp in length. The full length of the ORF was 1 635 bp, encoding 544 amino acid polypeptides. The value of the molecular weight was 59.63 ku, and the value of the isoelectric point was 9.02. By using RT-qPCR, the expression law of 4CL in 8 developmental stages of the V.amurensis coloration was analyzed. The gene expression of 4CL showed a trend of "low-high-low". The peak value was reached in the pericap full-color(100%) period, while a downward trend was showed in the full ripening stage. The prokaryotic expression recombinant plasmid pET28 a-4 CL was constructed and 4CL was expressed in E.coli BL21, which is consistent with the expected size.

关 键 词：山葡萄 果皮着色 4CL基因 原核表达 表达特征 

分 类 号：Q946.83[生物学—植物学]