山羊MSTN基因打靶载体的构建与鉴定

Construction and identification of goat myostatin gene targeting vector

根据绵羊、猪、小鼠和牛的肌肉生长抑制素(MSTN)基因序列设计引物,通过PCR方法扩增了山羊MSTN基因的5'同源臂和3'同源臂,其长度分别为5.2 kb、1.1 kb。在ploxPneo载体的neo基因上游和下游分别插入上述2个同源臂,经PCR、限制性内切酶鉴定及DNA序列测定,证实该载体的2条同源臂包含山羊MSTN基因的相应外显子及其邻近的部分内含子,从而成功构建了山羊ploxP-MSTN打靶载体。

Two pairs of primers were designed according to sequence of myostatin （ MSTN ） of sheep, pig, mouse and cattle. The 5＇ and 3＇ homologous arms of goat myostatin gene were amplified from the genomic DNA, which were about 5.2 kb and 1.1 kb in length, respec- tively. The two fragments were cloned into ploxPneo vector and located at both sides of the neomycin resistance gene. By PCR, restricted en- donucleases digestion and sequence analysis, the two homologous arms with partial corresponding exons and introns were demonstrated to be cloned into ploxPneo vector. In conclusion, the replacement targeting vector for goat MSTN gene has been successfully constructed.