体内精原干细胞转染法建立转基因小鼠

将人Bcl-2 cDNA与小鼠乳清酸蛋白(WAP)5’上游调控序列融合后，与脂质体按一定比例混合，再加入适量的台盼蓝制成转染液，注入到小鼠睾丸中的曲细精管中，转染精原干细胞以探讨建立转基因小鼠的可行性。共注射了3只公鼠，4天后将公鼠与发情母鼠合笼交配。共生仔鼠20只。检测结果表明，有3只呈PCR阳性，Southern blot检测，阳性鼠2只，1只公鼠，1只母鼠，其中，公鼠意外死亡；Western blot证实，1只母鼠的乳腺组织表达了Bcl-2蛋白，其F1代的16只小鼠中。有7只呈PCR阳性。证实了体内精原干细胞转染建立转基因动物的可行性。

Production of transgenic mice by in vivo spermatogonia-mediated gene transfer]

The present study examined if injection of DNA into the testis tabular could generate transgenic mice via transfecting spermatogonia. The 0.9 kb Bcl-2 cDNA was fused to the promoter region of mouse whey acid protein gene and the SV40 polyA. A 3.0 kb fragment of WAP-Bcl-2-SV40 was mixed with cationic liposome and one side tabular of 3 mice testis was injected with the fragment, the other side was ligatured. Two out of the 3 males were used to mate with the female 4 days later. Twenty pups were produced and 3 of which were proven to be gene integration positive by PCR detection. Two, 1 male and 1 female, were further confirmed to carry the transgene by Southern blot analysis. The male died by accident during its feeding. The female was demonstrated to express Bcl-2 protein in its mammary glands by Western blot assay. Seven out of 45 F1 mice were proven to be transgenic by PCR. It is concluded that transfecting spermatogonia in vivo can produce transgenic mice.