myostatin基因打靶的成肌细胞制备

Myostatin(MSTN), β-转化生长因子超家族的一员，是肌肉生长的负调控因子，该基因自然突变的比利时蓝牛和皮尔蒙特牛出现双肌性状。基因敲除该基因，是实现家畜产肉量的提高的有效方法。通过建立无启动子打靶载体MSTN-GFP, MSTN-neo，转染新生和胎儿绵羊骨骼肌细胞，经过表达绿色荧光蛋白报告基因和G418筛选获得骨骼肌细胞克隆，PCR，Southern blot,DNA序列检测获得阳性细胞克隆，为制备myostatin基因敲除的体细胞克隆绵羊提供核移植供体。

The Construct of Myostatin Gene Targeted Recipient Myoblast Cell

Abstract Myostatin (MSTN), a member of the transforming growth factor-β（TGF-β）superfamily, has been shown to be a negative regulator of myogenesis. Natural mutation in beef cattle causes double-muscling phenotypes. An investigation designed to knock out the myostatin gene by gene targeting in ovine myoblast cells were reported. Two promoter-trap targeting vectors MSTN-GFP and MSTN-Neo were constructed and used to transfect fetal and neonatal ovine primary myoblast cells. Both GFP expressing cells and drug resistant cells were obtained. Targeted cells expressing GFP were confirmed by PCR assay and drug resistant cells were characterized by PCR, Southern blot and DNA sequnce detection after growing into cell clones. That would supply nuleus transplantation donator for myostatin gene targetted somatic cloning sheep.