人c-myc乳腺定位表达转基因小鼠的建立

Establishment of Transgenic Mice Expressing c-myc Gene in Mammary Gland

以含有 1.6 kb的小鼠乳清酸蛋白 (WAP)上游调序列的 p CAT- WAP和含有人 c- myc c DNA的 p WM为原始质粒 ,构建了 WAP启动子调控下的 c- myc乳腺定位表达载体 p WCS。载体用 Bgl +Bam H 酶切 ,回收 3.5 kb的基因片段 WAP- c- myc- SV40 Poly A,通过显微注射方法导入 C5 7BL/ 6 J× DBA/ 2 JF1 代小鼠受精卵的雄原核内。共注射10 0 0枚卵 ,将存活的约 6 0 0枚卵分别移植至 2 9只假孕母鼠输卵管内 ,获得仔鼠 45只。PCR检测 ,阳性鼠 9只 ,South-ern blot检测 ,阳性鼠 3只 ,其中 1只母鼠 ,2只公鼠 ,在饲养过程中 ,1只母鼠意外死亡。对 2只转基因公鼠的 F1代PCR检测表明 ,仅 1只公鼠具有遗传性 ,在所生的 2 7只 F1代小鼠中有 13只为 PCR阳性。

c-myc cDNA was cloned on the downstream of mouse WAP promoter to form a expression vector pWCS.Transgenic mice were established by microinjection of the 3.5 kb c-myc cDNA expression cassette purified from pWCS.600 ova were transplanted into twenty-nine pseudopregnant mouse.recipients.45 offspring were born from the ova mircroinjected.9 of them were proved to be gene integration positively by PCR,3 mice,1 male and 2 female,were further confirmed to be transgenic by Southern hybridization.The female mice died by accidently during its feeding.1 of the other 2 male mice could inherit the c-myc gene into its F1 generation proved by PCR.6 female of 35 mice in its F1 generation were comfirmed to be transgenic by PCR.They are at a stage of mutiple pregnancy and lactation.