CAJ | 학술논문

目的：制备与鉴定神经干细胞特异性PPARγ基因敲除小鼠模型。方法将引进的2种转基因小鼠B6.PPARγloxp/loxp、B6. Nestin-Cre进行饲养并杂交繁殖，将子一代小鼠与B6.PPARγloxp/loxp小鼠回交获得子二代小鼠，提取子二代小鼠的基因组DNA，利用PCR方法扩增Cre和loxp基因片段，并进行琼脂糖凝胶电泳检测。选取基因型为B6.PPARγloxp/loxp.Nestin-Cre （KO）的小鼠即为神经干细胞特异性敲除PPARγ的敲除小鼠，另选基因型为B6.PPARγloxp/loxp（loxp）作为对照组小鼠。应用RT-PCR、实时荧光定量PCR方法鉴定神经干细胞特异性敲除PPARγ的敲除小鼠。结果敲除小鼠在基因鉴定时可以扩增得到PPARγloxp和Cre两个条带，在mRNA表型检测时脑内PPARγ表达显著低于对照组小鼠。成功获得神经干细胞敲除PPARγ基因的敲除小鼠。所购2种转基因小鼠均有繁殖能力，其繁殖符合孟德尔遗传规律。结论基于loxp-Cre系统成功构建神经干细胞特异性敲除PPARγ的基因敲除小鼠，为进一步的神经系统疾病的治疗及其机制研究提供模型基础。
목적：제비여감정신경간세포특이성PPARγ기인고제소서모형。방법장인진적2충전기인소서B6.PPARγloxp/loxp、B6. Nestin-Cre진행사양병잡교번식，장자일대소서여B6.PPARγloxp/loxp소서회교획득자이대소서，제취자이대소서적기인조DNA，이용PCR방법확증Cre화loxp기인편단，병진행경지당응효전영검측。선취기인형위B6.PPARγloxp/loxp.Nestin-Cre （KO）적소서즉위신경간세포특이성고제PPARγ적고제소서，령선기인형위B6.PPARγloxp/loxp（loxp）작위대조조소서。응용RT-PCR、실시형광정량PCR방법감정신경간세포특이성고제PPARγ적고제소서。결과고제소서재기인감정시가이확증득도PPARγloxp화Cre량개조대，재mRNA표형검측시뇌내PPARγ표체현저저우대조조소서。성공획득신경간세포고제PPARγ기인적고제소서。소구2충전기인소서균유번식능력，기번식부합맹덕이유전규률。결론기우loxp-Cre계통성공구건신경간세포특이성고제PPARγ적기인고제소서，위진일보적신경계통질병적치료급기궤제연구제공모형기출。
Objective To breed neual stem cell-specific peroxisome proliferator-activated receptor γ (PPARγ) knockout mice. Methods Two transgenic mouse models, namely B6.PPARγloxp/loxp and B6.Nestin-Cre were interbred, and the first-generation offsprings were backcrossed with B6.PPARγloxp/loxp to obtain the second-generation mice. Genomic DNA was extracted from the second-generation mice for PCR to amplify the loxp and Cre gene fragments followed by agarose gel electrophoresis to verify their sizes. The mice with the PPARγloxp/loxp.Nestin-Cre (KO) genotype were selected as the neural stem cell-specific knockout PPARγ mice, with B6.PPARγloxp/loxp (loxp) mice as the control. Tissue samples were collected from specific regions of the mouse brain and peripheral tissue for detecting the expression of PPARγ mRNA using RT-PCR and real-time quantitative PCR. Results and Conclusion Genotyping results showed PPARγloxp and Cre bands in the knockout mice, which showed obviously decreased mRNA expression of PPARγ, suggesting successful establishment of neural stem cell-specific PPARγ knockout mice. The two transgenic mice we used were fertile, and their breeding pattern followed the laws of Mendelian inheritance.