狗獾粪便DNA提取方法的初步探讨

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摘要

在京杭大运河扬州段堤坝上采集狗獾的新鲜粪便,采用预处理-酚/氯仿抽提法、NaCl改良法、异硫氰酸胍裂解法、淀粉吸附法及QIAamp试剂盒法5种方法对粪便样品中狗獾DNA进行提取,探讨狗獾粪便样品中DNA提取方法和优化条件。结果表明,在5种提取方法中,淀粉吸附法的效果明显优于其它4种方法。采用粪便裂解液快速裂解细胞后,加入淀粉去除其中的大量PCR抑制物,然后用蛋白酶K裂解、酚/氯仿/异戊醇抽提,最后使用UNIQ-10柱纯化粪便DNA,对线粒体控制区和微卫星位点的PCR扩增反应及测序结果证实该方法的可行性。以上结果表明,通过该方法获得的粪便DNA能够用于更深入的分子遗传学等学科的研究。

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	刘宇庆
	吕慎金
	殷宝法
	范红梅
	刘燕
	魏万红

关键词 ：狗獾, 粪便DNA, 提取, PCR扩增

Abstract：

The fresh feces of badger (Meles meles) were collected from the dam of Jing-Hang Canal near Yangzhou,and five DNA extraction methods including amylum-subsiding method,preprocess-organicsolvent extraction method,NaCl modified method,GuSCN-SiO₂ method and QIAamp DNA Stool Mini Kit method were used to extract the DNA from them.The results showed that the highest amplification success rate was in the amylum-subsiding method.In this study,fecal sample was lysed rapidly with digested solution,then DNA was extracted using chloroform/phenol after incubation with proteinase K,following by purification to remove numerous potential PCR inhibitors.The mitochondrial DNA control region and 5 microsatellite loci were amplified.showing that the mitochondrial DNA control region sequenced successfully.Our results suggested that DNA extraction based on this method was reliable.

Key words： badger(Meles meles) faecal DNA extraction PCR amplification

收稿日期: 2008-12-25

Q523+.8

基金资助:国家自然科学基金(30570289)

引用本文:

刘宇庆1,2, 吕慎金3, 殷宝法1, 范红梅1, 刘燕2, 魏万红1*. 狗獾粪便DNA提取方法的初步探讨[J]. , 2008, 27(6): 468-473.
LIU Yu-qing1,2, LV Shen-jin3, YIN Bao-fa1, FAN Hong-mei1, LIU Yan2, WEI Wan-hong1. A preliminary study on the DNA extraction method from faeces of badger (Meles meles). , 2008, 27(6): 468-473.

链接本文:

http://www.ecolsci.com/CN/Y2008/V27/I6/468

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