针对微量细胞样本进行细胞核和线粒体基因组同步快速分离的方法研究  

The Method of Simultaneous and Rapid Separation of Nuclear and Mitochondrial Genomes from Microcell Samples

作　　者：胡胜 胡哲[2] 汪方奎[2] Hu Sheng;Hu Zhe;Wang Fangkui(National Engineering Laboratory of On Site Physical Evidence Traceability Technology,Key Laboratory of Forensic Genetics,Ministry of Public Security,IFSC,Beijing 100038,China;National Key Laboratory of Agricultural Microbiology,Huazhong Agricultural University,Wuhan 430070,China)

机构地区：[1]公安部鉴定中心现场物证溯源技术国家工程实验室法医遗传学公安部重点实验室,北京100038 [2]华中农业大学农业微生物资源发掘与利用全国重点实验室,湖北武汉430070

出　　处：《山东化工》2024年第3期151-154,共4页Shandong Chemical Industry

基　　金：公安部鉴定中心开放课题(2019FGKFKT06)。

摘　　要：开发了一种针对微量细胞样本进行细胞核和线粒体基因组同步快速分离的方法。该方法首先利用体积分数1%非离子型表面活性剂NP-40能够选择性破坏细胞膜而维持细胞核完整性的特征,实现了细胞的轻柔裂解和细胞核完整性的维持;其次通过差速离心,实现细胞核(核基因组)与线粒体基因组的同步分离。经过显微成像观察,核酸蛋白分析仪检测以及PCR分析,实现了从10^(2)到10^(6)个悬浮培养细胞中高质量线粒体基因组的分离纯化。本方法获取的线粒体基因组无核基因组污染,为后续开展线粒体群体遗传分析、线粒体疾病诊断提供了一种高效、快速、低成本的解决方案。In this study,we developed a method for simultaneous and rapid separation of nuclear and mitochondrial genomes for microcell samples.In this method,the 1%nonionic surfactant NP40 can selectively destroy the cell membrane and maintain the nuclear integrity,so as to achieve the gentle lysis of cells and the maintenance of nuclear integrity.Secondly,by differential centrifugation,the nuclear genome and mitochondrial genome are separated synchronously.Through microscopic imaging observation,nucleic acid protein analyzer detection,and PCR analysis,we achieved the isolation and purification of high-quality mitochondrial genomes from 10^(2) to 10^(6) suspension culture cells.The mitochondrial genome obtained by this method is free of nuclear genome contamination,which provides an efficient,rapid and low-cost solution for subsequent mitochondrial population genetic analysis and mitochondrial disease diagnosis.

关 键 词：线粒体分离纯化 微量细胞 NP40缓冲液 

分 类 号：O65[理学—分析化学] Q2-33[理学—化学]