dspp-LacZ转基因小鼠G0代的制备和鉴定

Generation of dspp-LacZ transgenic mouse founders

目的:构建小鼠牙本质涎磷蛋白(DSPP)特异性启动子启动LacZ表达的转基因小鼠G0代。方法:用PCR方法获得DSPP编码序列上游约1.6kb的启动子序列,测序确认后,通过亚克隆方法将其与LacZ编码序列连接到同一个质粒中,构建转基因载体pTN-DPM-LacZ;将线性化的载体DNA注射到小鼠受精卵的雄原核,移植到假孕母鼠的输卵管;仔鼠出生后4周,用PCR检测阳性小鼠。结果:注射的受精卵共503个,移卵后产仔89只,阳性12只,阳性鼠分别传代开始建系。结论:通过显微注射方法获得12只G0代dspp-LacZ转基因小鼠,正在建立中的转基因小鼠系将来可以用作研究DSPP确切表达谱的工具。

PURPOSE: To establish a transgenic mouse founders in which the expression of LacZ was directed by a dentin sialophosphoprotein-specific promoter. METHODS: The DSPP-specific promoter was obtained by PCR and confirmed by sequencing, and the transgenic plasmid, pTN-DPM-LacZ, was constructed by subcloning the DSPP-specific promoter and LacZ-encoding sequence into one vector. The linearized transgenic plasmid was microinjected into the male pronucleus of the zygotes, and the microinjected zygotes were implanted to recipient pseudopregnant mice. The tail DNA of 4 week-pups was tested by PCR. RESULTS: 503 embryos were implanted to 20 recipient pseudopregnant mice, 12 of the 89 pups carrying the transgene. The establishment of the dspp-LacZ transgenic mouse line is under progress. CONCLUSION: 12 founders of the dspp-LacZ transgenic mice, in which the expression profile of LacZ should be the same as that of DSPP, were obtained by microinjection successfully, and the mouse line which is being established could be used as a good tool to investigate the exact expression profile of DSPP in the future.