小鼠胚胎的快速DNA提取及PCR性别鉴定

Rapid DNA extraction and PCR-sexing of mouse embryos.

作者信息

McClive P J, Sinclair A H

机构信息

Department of Paediatrics, University of Melbourne and Murdoch Children's Research Institute, Royal Children's Hospital, Melbourne, Parkville, Australia.

出版信息

Mol Reprod Dev. 2001 Oct;60(2):225-6. doi: 10.1002/mrd.1081.

DOI:10.1002/mrd.1081

PMID:11553922

Abstract

We have devised a PCR-based sexing method that is quick, simple, and highly reproducible. DNA is first extracted from embryonic mouse yolk sac via a 15 min, two-step incubation procedure utilizing PCR-compatible proteinase K buffer. Without any further manipulation the lysate is subjected to 30 cycles of PCR, optimized to run in less than 1 hr. The reaction includes multiplexed primer pairs for Sry and Myog (myogenin) that generate a male specific band of 441 bp and an internal control band of 245 bp, respectively. This robust method is used routinely in our laboratory and gives rapid genotyping results with 98% reliability and 100% accuracy.

摘要

我们设计了一种基于聚合酶链反应（PCR）的性别鉴定方法，该方法快速、简单且具有高度可重复性。首先通过使用与PCR兼容的蛋白酶K缓冲液进行15分钟的两步孵育程序，从胚胎小鼠卵黄囊中提取DNA。无需任何进一步处理，将裂解物进行30个循环的PCR，优化后运行时间不到1小时。该反应包括用于Sry和Myog（肌细胞生成素）的多重引物对，分别产生441 bp的雄性特异性条带和245 bp的内部对照条带。这种可靠的方法在我们实验室中常规使用，能快速给出基因分型结果，可靠性为98%，准确率为100%。