WSSV、IHHNV、EHP和NHPB四重荧光定量PCR检测方法的建立

Establishment of Quadruple-Fluorescence Quantitative PCR Detection Method for WSSV,IHHNV,EHP and NHPB

本研究以对虾白斑综合症病毒(White Spot Syndrome Virus,WSSV)的VP664基因、传染性皮下及造血组织坏死病毒(Infectious Subcutaneous and Hematopoietic Necrosis Virus,IHHNV)的NSP2基因、虾肝肠胞虫(Enterocytozoon Hepatopenaei,EHP)的ssr RNA基因以及坏死性肝胰腺炎细菌(Necrotizing Hepatopancreatitis Bacteria,NHPB)的16Sr RNA基因为检测对象,建立了WSSV、IHHNV、EHP和NHPB的四重荧光定量PCR方法,并对该方法的特异性、灵敏性进行了研究。结果表明,建立的四重荧光定量PCR方法特异性良好,对WSSV、IHHNV、EHP和NHPB的最低检测限为4.05 copies·μL^(-1)、6.53 copies·μL^(-1)、5.85 copies·μL^(-1)和6.18 copies·μL^(-1),本实验建立的方法具有较好的应用前景。

This study focuses on the VP664 gene of WSSV in shrimp,the NSP2 gene of IHHNV,the ssrRNA gene of EHP,and the 16SrRNA gene of NHPB.A quadruple fluorescence quantitative PCR method was established to simultaneously detect WSSV,IHHNV,EHP,and NHPB,and the specificity and sensitivity of this method were studied.The experimental results indicate that the established quadruple fluorescence quantitative PCR method has good specificity,with the minimum detection limits for WSSV,IHHNV,EHP,and NHPB being 4.05 copies·μL^(-1),6.53 copies·μL^(-1),5.85 copies·μL^(-1) and 6.18 copies·μL^(-1).The method established in this experiment has good application prospects.