蜜蜂球囊菌全长cDNA文库的构建与分析  

Construction and Analysis of a Full-length cDNA Library of Ascosphaera apis

作　　者：赵红霞[1,2] 李江红[1] 梁勤[1] 罗岳雄[2] 张学锋[2] 

机构地区：[1]福建农林大学蜂学学院,福州350002 [2]广东省昆虫研究所,广州510260

出　　处：《中国畜牧兽医》2010年第3期112-115,共4页China Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金项目(30600454);福建省自然科学基金项目(B0510023);蜂产业体系项目(nyhyzx07-041)资助

摘　　要：本试验利用TRIzol试剂盒提取蜜蜂球囊菌总RNA,通过Oligotex纯化试剂盒将mRNA反转录成cDNA,再以第一链cDNA为模板合成第二链cDNA,该cDNA产物经分级分离和体外包装,即获得蜜蜂球囊菌cDNA原始文库,其滴度测定为1.6×106PFU/mL,扩增后的滴度是1.5×109PFU/mL;取适量扩增文库稀释并铺平板,蓝白斑筛选独立噬菌斑,用M13±通用引物进行PCR扩增后,文库的重组率达到90%,插入cDNA片段的长度在0.5-2 kb。该文库的构建,有利于筛选目的基因,便于深入研究蜜蜂白垩病的侵染机制,最终有效防制白垩病。In this study,the total RNA of Ascosphaera apis was extracted by using TRIzol reagent.The Oligo reverse transcriptase was used to synthesize and anchor the first strand Cdna.Following the production of the first reverse transciption,we synthesize the second Cdna.The production of cDNA was separated by classes and packed outside,then we could get the cDNA original library of Ascosphaera apis.The unamplified cDNA library contaied 1.6×106 PFU/mL independent clones.The titer of the amplified library was 1.5×109 PFU/mL.Taking suitable amplified library to dilute and litter the flat,picking single plaques and then using common primer M13± to amplify,the recombination clones with an average insert size of 0.5 to 2 kb was higher than 90%.The construction of this library will provide a platform for the research of invasion mechanism and prevention.

关 键 词：蜜蜂球囊菌 全长CDNA文库 噬菌体 

分 类 号：S826.92[农业科学—畜牧学]