牛肌肉生成抑制素基因功能区的克隆与原核表达

Cloning and prokaryotic expression of bovine myostatin gene

根据GenBank中牛肌肉生成抑制素(MSTN)基因序列设计了1对引物,在引物两端分别加EcoR I和Xho I识别位点。利用RT-PCR技术扩增出了牛MSTN功能区序列。分别构建克隆和原核表达载体,酶切、PCR鉴定及测序分析表明,该基因功能区序列的克隆载体和原核表达载体已成功构建。筛选阳性菌,经IPTG诱导,牛MSTN基因功能区在大肠埃希氏菌中成功表达。

In order to express the functional sequence of bovine myostatin gene in the prokaryotic system, a pair of primers was designed according to the myostatin cDNA sequence and optimized with restriction sites for two restriction endonucleases EcoR Ⅰ and Xho Ⅰ . The functional fragment was amplified from bovine total RNA using RT-PCR. The cloning plasmid and pET-28a vector were digested, retrieved and sequenced, respectively. Double digestion with EcoR Ⅰ and Xho Ⅰ , PCR identification and sequence analysis showed that both the cloning vector and the prokaryotic expression vector for the functional fragment of the myostatin gene were constructed successfully. Positive clones were selected and induced by IPTG, and the expressed product of the bovine functional fragment was obtained.