摘要
为研究小鼠PTA1分子在体内的功能,建立四环素调控的小鼠PTA1/CD226转基因小鼠,我们构建了pBI-5-mPTA1载体,显微注射入B6D1F1受精卵,使用PCR检测新生小鼠基因组DNA中的PTA1与荧光素酶(luciferase)基因。将mPTA1和荧光素酶双阳性小鼠耳成纤维细胞转染含rtTA的pUHD17.1质粒,用含有盐酸强力霉素(Dox)的培养基进行培养,检测细胞裂解液中荧光素酶的活性。将荧光素酶表达依赖Dox的小鼠与C57BL/6小鼠交配,采用PCR对子代鼠进行检测。最终共获得7只首建鼠,其目的基因表达高度依赖Dox,并得到了其中2只首建鼠的F1代小鼠。
To establish the conditional tetracycline-regulated transgenic mice over-expressing mouse platelet and T cell activation antigen I（mPTA1）, the mPTA1 expression vector was constructed by recombination of pcDNA3- mPTA1 and pBI-5, and the transgenic mouse lines were generated by pronuclear injection using standard techniques. Positive animals were selected based on analysis of DNA sequence from mouse tails by PCR. The controlled regulation of the integrated expression unit was tested by transfection of the primary mouse-ear fibroblasts obtained from DNA-positive founders with the rtTA plasmid pUHD-17.1. Founder animals that showed Dox-dependent expression of luciferase were selected and interbred with C57BL/6 mice to establish a defined genetic background. We had created the transgenic mice, in which the expression of both mPTA1 and luciferase eDNA was found to be controlled by bidirectional promoter Pbi-1 Among 13 founder animals carrying the mPTA1 transgene, 7 founders could regulate luciferase in response to Dox as monitored in primary fibroblast cultures. Two lines were established after interbreeding with C57BL/6 mice. The conditional tetracycline-regulated transgenic mice over-expressing mPTA1 were established successfully, which may provide an useful animal model for investigating the function of mPTA1 molecule in vivo.
出处
《现代免疫学》
CAS
CSCD
北大核心
2005年第6期489-492,共4页
Current Immunology
基金
国家重点基础研究发展规划资助项目(2001CB510004)
