3种DNA提取方法对WSSV检测结果的影响  

The comparison of DNA extraction methods in White Spot Syndrome Virus detection

作　　者：李战军[1,2] 孔杰[2] 孟宪红[2] 张庆文[2] 罗坤[2] 栾生[2] 肖广侠[2,3] 

机构地区：[1]大连海洋大学生命科学与技术学院,116023 [2]农业部海洋渔业资源可持续利用重点开放实验室中国水产科学研究院黄海水产研究所,青岛266071 [3]上海海洋大学水产与生命学院,201306

出　　处：《渔业科学进展》2011年第3期92-97,共6页Progress in Fishery Sciences

基　　金：国家863计划项目(2010AA10A401);山东省自然科学基金项目(ZR2009DQ002);公益性事业(农业)科研专项(200803012);中央级公益性科研院所基本科研业务费2009年专项资金项目共同资助

摘　　要：比较了酚-氯仿法、煮沸法、试剂盒法3种基因组DNA提取方法对WSSV DNA提取效率、纯度及病毒检测结果的影响。结果表明,3种方法的DNA提取效率平均值分别为101.5、372.6、21.5 ng/μl;OD260/OD280范围分别是1.979~2.175（平均值为2.070）、1.699~1.932（平均值为1.796）、1.784~2.075（平均值为1.951）;OIE巢式PCR阳性率分别为60%、50%、70%;TaqMan定量PCR检测的病毒阳性率均为100%,病毒拷贝含量分别为916.0~2.23×106、63.3~1.78×106、479.7~2.70×106Copies/μl DNA。In this study,three genomic DNA extraction methods including phenol-chloroform extraction,boiling lysis and reagent kit methods were compared in terms of extraction efficiency,DNA quantity,as well as the results of White Spot Syndrome Virus（WSSV） detection.The DNA extraction efficiency of three methods,was 101.5,372.6 and 21.5 ng/μl,respectively.The ratios of OD260/OD280 were in the range of 1.979~2.175（mean 2.070）,1.699~1.932（mean 1.796） and 1.784~2.075（mean 1.951）,respectively.The positive rate of WSSV by OIE（Office International Des Epizooties） nested-PCR was 60%（DNA from phenol-chloroform extraction method）,50%（DNA from boiling lysis method） and 70%（DNA from kit method）,while it was 100% by quantitative PCR（all DNA from three methods）.The DNA copy number of WSSV detected by quantitative PCR ranged from 916.0 to 2.23×106,63.3 to 1.78×106 and 479.7 to 2.70×106 copies/μl,respectively.The results would provide some reference for standardizing the detection of WSSV.

关 键 词：DNA提取方法 PCR WSSV 

分 类 号：S962.32[农业科学—水产养殖]