模拟感染家蚕微粒子病的蚕卵、蚕蛾PCR检测的初步研究  

Preliminary Studies on the Application of PCR Diagnosis on the Silkworms Eggs and Moths With Simulated Pebrine Disease Infection

作　　者：刘吉平[1] 曹阳[1] Smith J E 徐兴耀[1] 

机构地区：[1]华南农业大学蚕学分子生物学与生物技术实验室,广州510642 [2]利兹大学生物学院

出　　处：《蚕业科学》2004年第4期367-370,共4页ACTA SERICOLOGICA SINICA

基　　金：英国DIFD资助项目 (编号CTN992 2 5 3 );广东省科技攻关项目 (编号C5 0 2 0 9)

摘　　要：在比较研究不同浓度家蚕微孢子虫 (Nosemabombycis,N .b)孢子与模拟染毒N .b孢子蚕卵、蚕蛾的模板DNA制备方法的基础上 ,选用MP1/MP2和V1F/ 5 30R两对引物进行PCR扩增检测。结果为 :碱性条件预处理N .b孢子后再抽提的DNA ,其得率略高于常规方法抽提的DNA ,但两者的模板质量相同 ;MP1/MP2和V1F/ 5 30R两对引物均可有效地检出N .b孢子DNA ,前者的检测灵敏度为 1μL 3× 10 6mL-1以上浓度的N .b孢子DNA ,后者为 1μL 3× 10 5mL-1以上浓度 ;对不同浓度N .b孢子DNA与蚕蛾DNA混合后进行PCR检测 ,V1F/ 5 30R引物的检测灵敏度为1μL 3× 10 6mL-1N .b孢子DNA。Based on the comparative studying on the methods of extracting the template DNA from the different concentration of purified Nosema bombycis(N.b),and from the simulated silkworm eggs and moths infected with N.b,and two sets of primer V1F/530R and MP1/MP2 were selected for PCR diagnosis.The results were as follows:the harvest ratio of N.b template DNA under alkaline pre-treatment was a little higher than that from the routine methods,however the quality of template DNA was adequate for PCR.Both sets of primer of V1F/530R and MP1/MP2 were useful to identify N.b DNA. The sensitivity of PCR primer V1F/530R for purified N.b DNA was approximately 3×105 mL -1or more,while MP1/MP2 primer was 3×106 mL -1or more.In PCR identification of the simulated silkworm moths DNA mixed with different concentration of N.b DNA,the sensitivity of V1F/530R was 3×106 mL -1.

关 键 词：家蚕 微粒子病 PCR诊断 模拟感染 微孢子浓度 

分 类 号：S884.2[农业科学—特种经济动物饲养]