胚胎发育早期转基因整合的研究

Studies of Foreign Gene Integeation During Embryo Early Development

转基因动物的建立是一项复杂而艰苦的工作，在转基因胚移植受体前对其进行检测，无疑对转基因动物建立具有重要意义。使用小鼠乳清酸蛋白（WAP）控制下的人粒细胞激落刺激因子（G-CSF）基因为显微注射片段，采用PCR方法检测了转基因胚，在1、2和8细胞期的阳性率为100％、77．7％和44．4％。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，转基因的非整合胚不能扩增出特异性片段。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，为在大动物转基因的检测提供了新依据。

There are still some problems in transgenic animals. Gene transfer, for example,reamins a difficult and costly task for animals, the vectors carrying the gene codingfor the proteins of interest are of unpredictable efficiency. Therefore, it is important toidentify foreign gene integration in genomose before transferring fertilized eggs toreceptors, in order to increase efficiency of producing transgenic animal. In thes paper,the constrict that mice whey acid protein (WAP) gene promotor directs G-CSF genewas used to microinject fertilized eggs of mice. Fertilized eggs containing foreign genewere measured by using PCR method. The results showed that 100%, 77.7% and44.4% retentions of foreign gene were achieved in 1, 2 and 8 cell-stage, respectively.Two part homologous recombination fragments were constructed and coinjected in tofertilized eggs of mice. PCR amplification fragment went beyond this homologousrecombination area. If foreign gene could not integrate in to genomose, the fragmentof PCR amplification could not be produced during embryo developmen. The resultsshowed that the rationes of foreign gene integrated in to genomoes in 1, 2 and 8cell-stage were 11.1%, 55.5% and 44.4%, respectively. This method might provide usa way to screen transgenic eggs when we use embryo section technique in farm animal.