一种从大熊猫粪便中提取DNA的改进方法  

An improved protocol for DNA extraction from the faeces of the giant panda

作　　者：钟华[1] 赖旭龙[2] 魏荣平[3] 刘中来[1] 

机构地区：[1]华中师范大学生命科学学院,武汉430079 [2]中国地质大学地球科学学院,武汉430074 [3]中国保护大熊猫研究中心,四川卧龙623006

出　　处：《动物学报》2003年第5期670-674,共5页ACTA ZOOLOGICA SINICA

基　　金：国家自然科学基金 (4 0 172 0 0 5)资助~~

摘　　要：本研究描述一个改进的方法 ,使从大熊猫粪便中提取DNA用于PCR扩增变得更加容易。在粪便DNA的提取过程中采用一个新的预处理方法 ,将粪便用预冷的丙酮洗 2～ 3次 ,除去粪便中含有的大量PCR抑制物 ,然后用蛋白酶K裂解、酚 -氯仿抽提 ,能提取到纯度很高的DNA供PCR扩增。本实验PCR扩增了大熊猫脑源性神经营养因子 (BDNF)基因和线粒体细胞色素b基因片段 ,并进行测序分析 ,证实了提取的可靠性。对比本方法和未经丙酮预处理的方法提取的DNA进行PCR扩增 。An improved method that facilitates the extraction of PCR compatible faecal DNA from giant panda's faeces is described. The method involved a novel preprocessing step in DNA extraction. The faeces was washed two or three times with precooled acetone, which removed numerous potential PCR inhibitors, and then digested with proteinase K. The DNA was purified with phenol/chloroform. The faecal DNA obtained was sufficiently pure to support reliable amplification, and was applied as template DNA to amplify a portion of the giant panda brain derived neurotrophic factor (BDNF) gene and mitochondrial cytochrome b gene. The sequenced results of PCR products confirmed that the extracted DNA was from the giant panda. Comparison with the PCR products demonstrated that the faecal DNA extracted by the improved protocol was better than the faecal DNA extracted without acetone preprocessing..

关 键 词：大熊猫 粪便DNA 丙酮 DNA抽提 非损伤性取样 

分 类 号：Q95-33[生物学—动物学]