悬铃木方翅网蝽DNA提取及PCR检测

目的]探索并建立悬铃木方翅网蝽总DNA提取方法。方法]采用改良SDS法结合Silica吸附柱提取悬铃木方翅网蝽总DNA,通过分光光度法、琼脂糖凝胶电泳对获得的DNA进行浓度、纯度及质量检测。利用PCR技术,分别扩增悬铃木方翅网蝽线粒体基因细胞色素氧化酶亚基Ⅰ（COⅠ）和基因组28S基因,克隆测序。结果]悬铃木方翅网蝽总DNA浓度为52.8μg/ml,A260/A280为2.06,琼脂糖凝胶电泳条带清晰。PCR扩增产物条带清晰特异,经克隆并测序后表明为悬铃木方翅网蝽COⅠ和28S基因。结论]该方法可得到较纯净完整和扩增效果良好的DNA,能满足进一步分子生物学研究的需要。

Extraction and PCR Detection of Total DNA of Corythucha ciliate Say

Objective] The research aimed to explore and establish a method of extraction of the total DNA from Corythucha ciliate Say.Method] The improved SDS method in combinaiton with a Silica adsorption column were used to extract the total DNA from Corythucha ciliate Say.The extracted DNA was qualitiatively assessed for the density and purity through spectrophotometry and agarose gel electrophoresis.Further the mitochondrial gene Cytochrome Oxidase Subunit Ⅰ(COⅠ) and the 28S gene in Corythucha ciliate Say genome were amplified,respectively by PCR,followed by cloning and sequencing.Result] The results showed that the total DNA density of Corythucha ciliate Say was 52.8 μg/ml witha ratio of A260/A280=2.06.In addition,the agarose gel electrophoresis band was clear.The bands of amplification outcome through PCR were clear and specific.The identify of the cloned fragment of the two gene preducts was further confirmed to be the Corythucha ciliate Say's COⅠ and the 28S gene,respectively,after cloning and sequencing.Conclusion] Using the improved SDS method high quality genomic DNA for further amplification usage could be obtain.The technique could conform with the needs of the future research of molecular biology.