天祝白牦牛Y染色体性别决定基因(SRY)编码区的克隆和原核表达  

Cloning of the Encoding Region of Sex-determining Region on Y Chromosome Gene(SRY) in Tianzhu White Yak and Its Prokaryotic Expression

作　　者：裴杰[1] 阎萍[1] 厍睿[2] 程胜利[1] 冯瑞林[1] 梁春年[1] 郭宪[1] 曾玉峰[1] 包鹏甲[1] 朱新书[1] 

机构地区：[1]中国农业科学院兰州畜牧与兽药研究所,兰州730050 [2]西北民族大学生命科学与工程学院,兰州730030

出　　处：《农业生物技术学报》2009年第4期609-613,共5页Journal of Agricultural Biotechnology

基　　金：中央级公益性科研院所基本科研业务费专项资金项目(No.BRF070105)资助

摘　　要：从天祝白牦牛(Bos grunneins)的基因组DNA中扩增了Y染色体性别决定基因(SRY)编码区序列,将其克隆至pGEM-Teasy载体,天祝白牦牛SRY基因编码区长687bp,编码229个氨基酸(GenBank:FJ373272);对牦牛和奶牛(B. grunneins)的SRY基因编码区进行序列比对,发现存在2个碱基的变异,造成1个氨基酸的变异;将牦牛SRY基因编码区连接至pET-28a(+)载体,成功地构建了表达载体pET-28a/SRY;把表达载体pET-28a/SRY转入大肠杆菌(Escherichia coli)BL21(DE3)中,用IPTG在30℃200r/min条件下诱导,获得了SRY蛋白的大量表达;对表达产物进行Western blot检测,结果显示获得的蛋白能与抗-His单抗特异性结合,确定了牦牛SRY蛋白的表达。The encoding region of sex-determining region on Y chromosome（SRY） was amplified from genome in Tianzhu White Yak（Bos grunneins）. The product was cloned to pGEM-T easy vector and sequenced. The total coding region of yak＇ s SR Y gene was 687 bp, encoding a peptide with 229 amino acid residues （GenBank： FJ373272）. Sequence alignment showed that there were two mutations in the coding region compared with cow （B. taurus）, causing one amino acid mutation in LF protein. The coding region ofyak＇s SRYgene was cloned to EcoR I and Sal I sites ofpET-28a （＋） vector to construct an expression plasmid pET-28a/SRY, which was then transformed into Escherichia coli BL21 （DE3）, and the SRY protein was highly expressed under 30℃ 200 r/min induced by IPTG. Westem blotting results showed that the protein obtained could bind with anti-His tag, indicating that the SRY protein of yak was expressed.

关 键 词：牦牛 Y染色体性别决定基因(SRY) 克隆 表达 

分 类 号：S188[农业科学—农业基础科学]