In order to understand the action mechanism of yak MSTN, seek for possible method to block its activity and finally realize an efficient growth in yak, MSTN gene was recombinantly expressed in prokaryotic cell and purified for testing its immunogenicity. Firstly, yak MSTN mature peptide gene was cloned using specific primers and reverse-transcription PCR method. Secondly, the target fragment was cloned into pET28a( + ) vector and then transformed into E. coli BL21 (DE3) for prokaryotic expression. The target peptide was purified from total prokaryotic expression products. Finally, immunogenicity of yak MSTN peptide was detected by ELISA. The MSTN mature peptide gene inserted into the recombinant expression vector contained 330 bp and encoded 109 amino acids in yak. The expression host transformants were induced by adding 0. 1 mmol o L-1 IPTG to the LB medium, and the content of target protein was about 21% of the total proteins after six hours of induction. The highly purified yak MSTN mature peptide was obtained by affinity chromatography and its molecular weight was 16. 5 ku. The result of ELISA showed that the mature yak MSTN recombinant peptide had an excellent immunogenicity and the titer of rabbit serum anti-yak MSTN was 1 : 32 000. The study of highly efficient expressed and purified yak mature MSTN peptide with excellent immunogenicity obtained from this study paved a way to study its functional mechanism that may be further explored to improve the meat production performance of yak.%为了探索牦牛MSTN的作用机制,寻找合适的阻断MSTN活性的方法,最终实现牦牛的高效率育肥,本研究采用RT-PCR方法克隆牦牛MSTN成熟肽基因,构建MSTN-pET28a(+)重组表达载体,进而在E.coliBI21(DE3)宿主菌中进行表达,将表达产物用亲和层析纯化,最后用ELISA方法检测牦牛MSTN重组成熟肽的免疫原性.重组载体中连接的牦牛MS TN成熟肽基因包含330 bp,编码109个氨基酸.含MSTN-pET28a(+)重组载体的工程菌经0.1 mmol·L-1 IPTG诱导表达6h后,MSTN成熟肽的表达量约占蛋白总量的21％,总蛋白经亲和层析纯化后得到了高纯度的牦牛MSTN重组成熟肽,分子量约16.5 ku；ELISA结果显示,兔抗牦牛MSTN重组成熟肽的血清效价为32 000个抗体单位,表明牦牛MSTN重组成熟肽具有较好的免疫原性.本研究建立了牦牛MSTN重组成熟肽高效表达系统并纯化获得具有免疫原性的牦牛重组MSTN成熟肽,这为通过调控MSTN功能来改良牦牛产肉性能的研究储备了技术条件.
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