基于多重PCR的寡核苷酸基因芯片检测猪瘟病毒,猪繁殖与呼吸障碍综合征病毒及猪圆环病毒1型和2型  

A multiplex PCR based oligonucleotide microarray for detection and differentiation of classical swine fever virus,porcine reproductive and respiratory virus and porcine circovirus types 1 and 2

作　　者：姜永厚[1] 商晗武[2] 徐辉[3] 朱良俊[1] 丁先锋[1] 陈伟杰[3] 赵灵燕[3] 方立[4] 

机构地区：[1]浙江理工大学生命科学学院,浙江杭州310018 [2]中国计量学院生命科学学院,浙江杭州310018 [3]浙江省动物疫病预防控制中心,浙江杭州310020 [4]浙江大学动物科学学院,浙江杭州310029

出　　处：《中国预防兽医学报》2010年第3期194-199,共6页Chinese Journal of Preventive Veterinary Medicine

基　　金：浙江省科技计划项目(2008C22081);浙江省自然科学基金项目(Y3090166);浙江省科技攻关重点项目(2005C22032)

摘　　要：为了建立一种可同时检测区分猪瘟病毒(CSFV),猪繁殖与呼吸障碍综合征病毒(PRRSV)和猪圆环病毒2型(PCV-2)的寡核苷酸基因芯片方法,本研究针对每种病毒设计了4条～6条寡核苷酸探针,检测低限为1.6×104PCV-2拷贝数/μL,1.6×104CSFV拷贝数/μL,1.6×105PRRSV拷贝数/μL,比琼脂糖凝胶灵敏约10倍。利用建立的寡核苷酸基因芯片方法对76个仔猪样本进行了检测,检出了3种病毒的存在,其中25个样本(32.9%)同时感染了2种以上病毒。结果表明寡核苷酸基因芯片检测是一种快速、灵敏和高效的猪只混合感染病毒的病原学诊断方法。Porcine circovirus type 2 （PCV-2）,classical swine fever virus （CSFV） and porcine reproductive and respiratory syn-drome virus （PRRSV） are important viruses that cause severe reproductive and/or respiratory failure in swine,and rapid and accu-rate detection and identification of these pathogens is critical for swine disease management. In the present study,we developed a multiplex polymerase chain reaction （PCR） based oligonucleotide microarray assay that included four to six specific oligonu-cleotides for each virus. The microarray was tested on three target viruses and 76 clinical specimens. The detection limit of the oligonucleotide microarray was 1.6×104,1.6×104,1.6×105 copies/μL for PCV-2,CSFV and PRRSV,respectively,which was 10-fold more sensitive than that of agarose gel. Co-infection with two or three viruses was detected in 26 specimens （32.8 %）. Theresults indicated that the microarray detection system is a reliable and effective method for viral pathogen detection and identification even when multiple pathogens are present in a porcine clinical sample.

关 键 词：基因芯片 猪瘟病毒 猪繁殖与呼吸障碍综合征病毒 猪圆环病毒 

分 类 号：Q789[生物学—分子生物学]