In this study,a RT-PCR assay for rapid detection of duck hepatitis virus type 1( DHV-1) was established in the presence of a pair of primers designed based on vp1 gene sequence,using the genomic RNA of DHV-1 or other ...In this study,a RT-PCR assay for rapid detection of duck hepatitis virus type 1( DHV-1) was established in the presence of a pair of primers designed based on vp1 gene sequence,using the genomic RNA of DHV-1 or other common viruses as the template. The template was serially diluted 10-fold to determine the reaction sensitivity. Finally,seven liver samples of ducks with suspected DHV-1 infection,which were collected from different regions of Jiangsu Province,were detected with this method. The results showed that a DNA fragment of about 360 bp was specifically amplified with the RT-PCR system,and the detection limit was1 fg/μL. The detection rate of clinical samples with this method was 100%. The results revealed that the RT-PCR system which had high specificity and high sensitivity in the detection of DHV-1 was successfully established.展开更多
本研究以日本黄瓜绿斑驳花叶病毒(kyuri green mottle mosaic virus,KGMMV)为材料,设计1对常规RT-PCR检测引物KGCPN-F/KGCPN-R和1组引物/探针KGM-F/KGM-R/KGM-P,分析测试了2对已发表KGMMV常规RT-PCR检测引物KGCP-F/KGCP-R和KGMP-F/KGMP...本研究以日本黄瓜绿斑驳花叶病毒(kyuri green mottle mosaic virus,KGMMV)为材料,设计1对常规RT-PCR检测引物KGCPN-F/KGCPN-R和1组引物/探针KGM-F/KGM-R/KGM-P,分析测试了2对已发表KGMMV常规RT-PCR检测引物KGCP-F/KGCP-R和KGMP-F/KGMP-R,建立了KGMMV的常规和实时荧光RT-PCR检测方法。结果表明,引物KGCP-F/KGCP-R扩增KGMMV时出现预期大小的条带,扩增黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)时出现非常微弱的条带;引物KGCPN-F/KGCPN-R扩增KGMMV时出现预期大小的条带,扩增小西葫芦绿斑驳花叶病毒(zucchini green mottle mosaic virus,ZGMMV)时出现比预期稍大的条带,通过对PCR产物进行序列测定和分析比对可准确鉴定KGMMV。KGCP-F/KGCP-R和KGCPN-F/KGCPN-R的相对灵敏度分别为10^(-6)和10^(-5)稀释度,适用于KGMMV的常规RT-PCR检测。基于引物探针KGM-F/KGM-R/KGM-P建立的KGMMV实时荧光RT-PCR检测方法能特异性检出KGMMV,相对灵敏度达10^(-7)稀释度,分别比2对常规RT-PCR检测引物高10倍和100倍,适用于瓜类种子中KGMMV的快速检测。展开更多
基金Supported by Natural Science Research Program of the Higher Education Institutions of Jiangsu Province(16KJB230004)"Six Talent Peaks" Project of Jiangsu Province(NY-023,10410015002)+1 种基金Anhui Science and Technology Project(1201c0602006)Jiangsu Province Excellent Scientific and Technological Innovation Tean
文摘In this study,a RT-PCR assay for rapid detection of duck hepatitis virus type 1( DHV-1) was established in the presence of a pair of primers designed based on vp1 gene sequence,using the genomic RNA of DHV-1 or other common viruses as the template. The template was serially diluted 10-fold to determine the reaction sensitivity. Finally,seven liver samples of ducks with suspected DHV-1 infection,which were collected from different regions of Jiangsu Province,were detected with this method. The results showed that a DNA fragment of about 360 bp was specifically amplified with the RT-PCR system,and the detection limit was1 fg/μL. The detection rate of clinical samples with this method was 100%. The results revealed that the RT-PCR system which had high specificity and high sensitivity in the detection of DHV-1 was successfully established.
文摘本研究以日本黄瓜绿斑驳花叶病毒(kyuri green mottle mosaic virus,KGMMV)为材料,设计1对常规RT-PCR检测引物KGCPN-F/KGCPN-R和1组引物/探针KGM-F/KGM-R/KGM-P,分析测试了2对已发表KGMMV常规RT-PCR检测引物KGCP-F/KGCP-R和KGMP-F/KGMP-R,建立了KGMMV的常规和实时荧光RT-PCR检测方法。结果表明,引物KGCP-F/KGCP-R扩增KGMMV时出现预期大小的条带,扩增黄瓜绿斑驳花叶病毒(cucumber green mottle mosaic virus,CGMMV)时出现非常微弱的条带;引物KGCPN-F/KGCPN-R扩增KGMMV时出现预期大小的条带,扩增小西葫芦绿斑驳花叶病毒(zucchini green mottle mosaic virus,ZGMMV)时出现比预期稍大的条带,通过对PCR产物进行序列测定和分析比对可准确鉴定KGMMV。KGCP-F/KGCP-R和KGCPN-F/KGCPN-R的相对灵敏度分别为10^(-6)和10^(-5)稀释度,适用于KGMMV的常规RT-PCR检测。基于引物探针KGM-F/KGM-R/KGM-P建立的KGMMV实时荧光RT-PCR检测方法能特异性检出KGMMV,相对灵敏度达10^(-7)稀释度,分别比2对常规RT-PCR检测引物高10倍和100倍,适用于瓜类种子中KGMMV的快速检测。
