坦布苏病毒是新发现的一种可引起家禽产蛋和采食量下降及死亡的黄病毒属病毒。根据GenBank中登录的鹅源坦布苏病毒NS5基因序列,利用Primer Explorer V4软件在序列保守区域设计1套特异识别NS5基因序列中6个不同区段的环介导等温扩增(loop...坦布苏病毒是新发现的一种可引起家禽产蛋和采食量下降及死亡的黄病毒属病毒。根据GenBank中登录的鹅源坦布苏病毒NS5基因序列,利用Primer Explorer V4软件在序列保守区域设计1套特异识别NS5基因序列中6个不同区段的环介导等温扩增(loop-mediated isothermal amplification,LAMP)引物,并以此套引物建立一种基于LAMP技术的鹅源坦布苏病毒检测方法。结果表明,该方法灵敏度极高,是普通PCR方法的100倍,只能特异性扩增坦布苏病毒的RNA,对于其他常见禽RNA病毒均无特异性扩增。在反应体系中加入SYBR GreenⅠ染料后,通过肉眼观察有无绿色荧光可直接判定结果。该方法特异性强,灵敏度高,无需特殊仪器,简便,快速,适用于鹅源坦布苏病毒的临床快速检测。展开更多
Reverse transcription polymerase chain reaction (RT-PCR) was used for the detection of classical swine fever virus (CSFV) in blood and tissue samples of field cases and experimentally inoculated pigs. The distribution...Reverse transcription polymerase chain reaction (RT-PCR) was used for the detection of classical swine fever virus (CSFV) in blood and tissue samples of field cases and experimentally inoculated pigs. The distribution of CSFV in different organ samples showed some discrepancies in infected pigs. Four weaner pigs were inoculated with C-strain vaccine virus, then samples of spleen, tonsil, lung, mesenteric lymph node, kidney and brain were collected after slaughter and tested for E2 and NS5B genes using one-step RT-PCR and nested RT-PCR. Using the same method, 12 field cases were simultaneously studied. A discrepancy of CSFV in different samples was found upon detecting the target gene. The most reliable diagnostic organs were spleen and tonsil, and the nested RT-PCR assay provided a highly sensitive and specific method with comparable performance to the one-step RT-PCR assay.展开更多
对大理地区2014年流行的丙型肝炎病毒(Hepatitis C virus,HCV)的非结构蛋白5(Nonstructural protein 5,NS5)基因序列进行分析研究,以了解云南省大理地区流行的HCV基因型或基因亚型。2014年1~10月在大理州人民医院收集临床诊断丙型肝炎...对大理地区2014年流行的丙型肝炎病毒(Hepatitis C virus,HCV)的非结构蛋白5(Nonstructural protein 5,NS5)基因序列进行分析研究,以了解云南省大理地区流行的HCV基因型或基因亚型。2014年1~10月在大理州人民医院收集临床诊断丙型肝炎病人血清,共收集到血清48份;对这些标本进行病毒核酸提取和反转录合成cDNA,采用HCV NS5基因特异引物进行RT-PCR扩增,27份标本扩增阳性,序列分析结果显示:5株与基因3a型HCV位于同一进化簇中,核苷酸同源性在90.8%~98.3%之间;14株与基因3b型位于同一进化簇中,核苷酸同源性在87.5%~98.6%之间;5株与基因1b型HCV位于同一进化簇中,核苷酸同源性在91%~97.7%之间;3株与基因6n型位于同一进化簇中,核苷酸同源性在92.8%~96.5%之间,表明本次分离到的27株HCV病毒中,5株属于3a型,14株属于3b型,5株属于1b型,3株属于6n型等3个基因型4个基因亚型。以上研究提示大理地区至少存在1b、3a、3b和6n基因亚型HCV流行,但以基因3型(3a、3b)为主要流行HCV基因型或基因亚型。展开更多
基金supported by the grants from the National"863" Programme (2006AA10A2041)Development Programme of Science and Technology,Chengguan District of Gansu province (08-5-4)
文摘Reverse transcription polymerase chain reaction (RT-PCR) was used for the detection of classical swine fever virus (CSFV) in blood and tissue samples of field cases and experimentally inoculated pigs. The distribution of CSFV in different organ samples showed some discrepancies in infected pigs. Four weaner pigs were inoculated with C-strain vaccine virus, then samples of spleen, tonsil, lung, mesenteric lymph node, kidney and brain were collected after slaughter and tested for E2 and NS5B genes using one-step RT-PCR and nested RT-PCR. Using the same method, 12 field cases were simultaneously studied. A discrepancy of CSFV in different samples was found upon detecting the target gene. The most reliable diagnostic organs were spleen and tonsil, and the nested RT-PCR assay provided a highly sensitive and specific method with comparable performance to the one-step RT-PCR assay.
文摘对大理地区2014年流行的丙型肝炎病毒(Hepatitis C virus,HCV)的非结构蛋白5(Nonstructural protein 5,NS5)基因序列进行分析研究,以了解云南省大理地区流行的HCV基因型或基因亚型。2014年1~10月在大理州人民医院收集临床诊断丙型肝炎病人血清,共收集到血清48份;对这些标本进行病毒核酸提取和反转录合成cDNA,采用HCV NS5基因特异引物进行RT-PCR扩增,27份标本扩增阳性,序列分析结果显示:5株与基因3a型HCV位于同一进化簇中,核苷酸同源性在90.8%~98.3%之间;14株与基因3b型位于同一进化簇中,核苷酸同源性在87.5%~98.6%之间;5株与基因1b型HCV位于同一进化簇中,核苷酸同源性在91%~97.7%之间;3株与基因6n型位于同一进化簇中,核苷酸同源性在92.8%~96.5%之间,表明本次分离到的27株HCV病毒中,5株属于3a型,14株属于3b型,5株属于1b型,3株属于6n型等3个基因型4个基因亚型。以上研究提示大理地区至少存在1b、3a、3b和6n基因亚型HCV流行,但以基因3型(3a、3b)为主要流行HCV基因型或基因亚型。