优化整体原位杂交方法检测基因在幼鼠大脑中的表达  

Detection of mRNA Expression on Postnatal Mouse Brains by Optimized Whole Mount in situ Hybridization

作　　者：刘安军[1,2] 麻献华[1] 杨瑞[1] 章卫平[1] 谢志芳[1] 

机构地区：[1]中国人民解放军第二军医大学基础部,上海200433 [2]中国人民解放军济南军区总医院麻醉科,250031

出　　处：《医学研究杂志》2016年第5期50-53,共4页Journal of Medical Research

基　　金：国家自然科学基金资助项目(31470759;31270814)

摘　　要：目的优化整体原位杂交方法,用于检测基因在幼鼠大脑中的时空表达分布。方法通过摸索最佳蛋白酶K的消化时间,改良杂交缓冲液成分,并延长杂交时间和漂洗时间,对整体原位杂交方法进行优化,并用优化的方法检测了Cdh6、Bhlhb5和Lmo4 mRNA在出生后7天的小鼠大脑中的整体表达分布。结果采用优化的整体原位杂交方法成功地检测了3种mRNA在幼鼠全脑的区域性表达特征,特异性杂交信号强,非特异性背景低。结论成功优化了幼鼠全脑原位杂交方法,为开展后续的研究打下基础。Objective To optimize whole mount in situ hybridization protocol for detecting mRNA expression on postnatal mouse brains. Methods Critical steps of whole mount in situ hybridization were optimized including proteinase K digestion time, composition of hybridization buffer as well as durations of incubation and washing steps. The mRNA expression of Cdh6, Bhlhb5 and Lmo4 was detected on brains from postnatal 7 days mice using the modified whole mount in situ hybridization protocol. Results By using the optimized whole mount in situ hybridization protocol, mRNA expression patterns of Cdh6, Bhlhb5 and Lmo4 were successfully detected on mouse brains, with great specificity and low background signal. Conclusion The optimized whole mount in situ hybridization protocol can be used for large -size organs such as postnatal mouse brains.

关 键 词：整体原位杂交 小鼠 大脑 基因表达 

分 类 号：R34[医药卫生—基础医学]