广西巴马小型猪卵泡抑素的克隆及原核表达  

Cloning and Prokaryotic Expression of Follistatin Gene from the Guangxi Bama Mini-pig

作　　者：莫毅[1] 郭亚芬[1] 梁方方 兰干球[1] 蒋和生[1] 李柏[1] 

机构地区：[1]广西大学动物科学技术学院,南宁530004 [2]广西畜牧研究所,南宁530001

出　　处：《中国畜牧兽医》2007年第10期49-52,共4页China Animal Husbandry & Veterinary Medicine

基　　金：广西教育厅基金项目

摘　　要：从广西大学巴马小型猪卵巢中提取总RNA,并以其为模板,应用逆转录-聚合酶链式反应(RT-PCR),在合成的特异性引物引导下,扩增获得广西巴马小型猪卵泡抑素(follistatin)cDNA的全序列,长度为1035 bp。将PCR产物克隆到pMD18-T载体后进行序列测定及分析,结果表明:广西巴马小型猪卵泡抑素cDNA序列与GenBank中已报道的家猪卵泡抑素同源性高达99.4%。同时,将目的基因插入到原核表达载体pET 32a+多克隆位点中,构建了follistatin的原核表达载体,并转化于大肠杆菌BL21。转化菌经异丙基硫代半乳糖苷(IPTG)诱导,变性聚丙烯酰胺凝胶电泳(SDS-PAGE)和蛋白质免疫印迹(Western blotting)分析,结果表明:试验已成功表达了follistatin的融合蛋白(约58.4 ku)。In this study, the total RNA was isolated from Guangxi Bama mini-pig ovary tissue with RNAwatson extraction. The complementary DNA （eDNA） encoding follistatin protein was ampilified by the reverse transcription polymerase chain reaction （RT-PCR） method and a pair of differential primers. The amplified eDNA fragment was inserted into pMD18-T vector. The recombined vectors were verified through the sequence survey. The results showed that the cloned follistatin eDNA was highly homologous with the reported nucleotide encoding swine follistatin in GenBank（99.4%）. Meanwhile, the identified follistatin eDNA was inserted in the multiple cloning sites of plasmid pET 32a＋ to construct prokaryotic expression vector, the latter was transformed into the E coli BL21 induced by IPTG. Then a fusion protein was obtained （about 58.4 ku） in SDS- PAGE and Western blotting. These results showed that the follistatin eDNA was expressed in prokaryotic cells.

关 键 词：广西巴马小型猪 卵泡抑素 克隆 原核表达 

分 类 号：Q786[生物学—分子生物学]