PCR扩增同源重组片段筛选转基因胚胎

使用小鼠乳清酸蛋白基因（ＷＡＰ）启动子控制下的人集落刺激因子（Ｇ－ＣＳＦ）基因为显微注射片段，采用ＰＣＲ方法检测了转基因胚，为消除ＰＣＲ扩增中的假阳性结果，构建了两个具有部分同源性的亚克隆片段进行共注射．ＰＣＲ扩增片段跨越这一同源区域，仅当注射的片段能够整合并发生正常重组，转基因整合胚才能以相对高的比例扩增出特异性片段．结果表明，１、２和８细胞期的阳性率分别为１１．１％、５５．５％和４４．４％，较常规ＰＣＲ检测获得更为明确的结论，为在大动物转基因胚胎检测提供了依据

Identification of Transgenic Embryo Using Homologous Recombinant Fragments

It is important to identify foreign gene integration in genomose before transferring fertilized eggs to receptors in order to increase efficiency of producing transgenic animals.In this paper,PCR method was used to identify transgenic embryos to screen foreign genes integrating and no integating embryos.The construction that mice whey acidic protein (WAP) gene 2 6 kb promoter directed human granulocyte colony stimulating factor(G CSF) gene was used to microinject fertilized eggs of mice.In order to decrease the mistakes caused by PCR in screening transgenic embryo,two part of homologous recombinant fragments were constructed and used for co microinjection in to fertilized eggs of mice.PCR amplification fragment went beyond this homologous recombinant area.If foreign gene could be integrated to genomose,and two fragments of co microinjection could create homologous recombinant in embryos,the fragment of PCR amplification in embryos could be produced during embryo development.The results showed that specific fragments of PCR amplification could be obtained in different embryo developments and foreign genes integrated to genomoes cell stages 1,2 and 8 were 11 1%,55 5% and 44 4%,resprectively.Compared with other methods to screen transgenic embryo,this method might provide us a way to screen transgenic eggs efficiently when we use embryo section technique in farm animals.