人bcl-xl转基因小鼠外源基因的复制及其传代稳定性

背景:近年的研究表明,在脑缺血再灌注损伤机制中,bcl-xl基因可能起着抗神经细胞凋亡的脑保护作用。目的:观察转bcl-xl基因小鼠外源基因的复制及传代稳定性问题。设计:以基因为观察对象。单位:华中科技大学同济医学院附属同济医院神经科和湖北省农科院畜牧兽医研究所。材料:实验于1999-10/2001-04在华中科技大学同济医学院附属同济医院完成。选取昆明种小鼠4只。方法:通过显微注射法建立转人bcl-xl基因小鼠,将PCR及Southern-blot检测整合阳性的小鼠与正常鼠交配并传代,然后检测子代小鼠外源基因的表达情况。作Southern-blot检测,结果证实第10,13,5,6号小鼠为整合阳性小鼠。以4只整合阳性小鼠作为首建者,分别与正常的昆明白小鼠交配繁殖传代,所生子代记为F1代,各系列的F1～F4代均采用阳性转基因小鼠与正常的昆明白小鼠交配繁殖传代方法,保持其杂合子形式。主要观察指标:检测外源基因的复制和传代的稳定性。结果:10号小鼠传代过程中外源基因PCR检测阳性率保持稳定,符合孟德尔遗传规律,表明外源基因能够稳定遗传。6号小鼠PCR阳性率呈轻微下降趋势,但仍能够保持相对稳定遗传。13号和5号小鼠PCR阳性率均呈明显的下降趋势,表明这2只小鼠存在外源基因遗传丢失现象。结论:应用基因组DNA做目的基因,可获得能够稳定遗传的转基因鼠系。

Reproduction and stable passage of foreign gene in human bcl-xl transgenic mice

BACKGROUND: Recent studies indicate that bcl-xl gene may inhibit the apoptosis of neurons in cerebral ischemia and repeff usion injury to exert the brain protection function.OBJECTIVE: To develop human bcl-xl transgenic mice by microinjection and detect the reproductionof the foreign gene and the stability of the passage.DESIGN: A gene-based observation.SETTINGS: Department of Neurology, Tongji Hospital affiliated to Tongji Medical College, Huazhong University of Science and Technology; Hubei Academy of Agriculture and Forestry Sciences, Research Institute of Animal Husbandry and Medicine.MATERIALS: The experiment was carried out in the Tongji Hospital affiliated to Tongji Medical College, Huazhong University of Science and Technology. Four Kunming mice were used.METHODS: Human bcl-xl gene transgenic mice were established with microinjection. The positive mice with integration detected by polymerase chain reaction (PCR) and Southern-blot were copulated with normal mice and passaged, and then the heredity of foreign gene in the filial generation was detected. The results of Southern-blot assay showed that the mice of No. 10, 13, 6 and 5 were positive, and they were taken as the initial ones, copulated with the normal Kunming mice respectively, proliferated and passed to the filial generation, called generation F1. The F1 to F4 of each series were produced by such a way to keep them heterozygotes.MAIN OUTCOME MEASURES: The reproduction and stable transmissibility of foreign gene in the offspring were detected.RFSULTS: The PCR positive rate of No. 10 mouse kept stable, accorded with the Mendelian inheritance rule, and the foreign gene could inherit steadily. The PCR positive rate of No.6 mouse showed the tendency of a little descent, but could maintain the relatively stable inheritance. For No.13 and 5 mice, there were notable descent, indicating that the foreign gene in this mice family had been lost during the heredity.CONCLUSION: Transgenic mice line can be obtained by taking genome DNA as the aim gene.