林业科学
林業科學
임업과학
SCIENTIA SILVAE SINICAE
2001年
3期
78-82
,共5页
赵同海%张永安%王玉珠%陈昌洁
趙同海%張永安%王玉珠%陳昌潔
조동해%장영안%왕옥주%진창길
松毛虫质型多角体病毒%多角体蛋白基因%反转录聚合酶链式反应
鬆毛蟲質型多角體病毒%多角體蛋白基因%反轉錄聚閤酶鏈式反應
송모충질형다각체병독%다각체단백기인%반전록취합매련식반응
为研究DsCPV在松毛虫种群中的垂直传递及对松毛虫灾害的持续控制作用，通过反转录聚合酶链式反应(RT-PCR)建立了DsCPV的早期检测技术。依据家蚕质型多角体病毒(BmCPV)质型多角体蛋白的核苷酸序列设计一对引物，从纯化的DsCPV\,BmCPV和舞毒蛾质型多角体病毒(LdCPV)的基因组dsRNA可成功地扩增出长614bp的目的片段，从提取的健康松毛虫幼虫肠组织的DNA、舞毒蛾核型多角体病毒(LdNPV)基因组核酸、以及棉铃虫质型多角体病毒(HaCPV)基因组核酸，未能扩增出目的片段。DsCPV基因组dsRNA扩增片段的序列与BmCPV相应基因序列具有87%的同源性，检测敏感度为1pg的DsCPV基因组dsRNA。由于松毛虫与家蚕、舞毒蛾相互之间食性不同，而且BmCPV和LdCPV对松毛虫无感染性，即在松毛虫体内不会有BmCPV病毒和LdCPV病毒的感染，因此该结果一方面从分子水平上证实了DsCPV与BmCPV、LdCPV存在基因同源性，同时也表明了依据BmCPV的基因序列设计引物对DsCPV基因组核酸建立的RT-PCR扩增体系，可以作为松毛虫种群中DsCPV的一种敏感、特异、早期、快速的检测手段。
為研究DsCPV在鬆毛蟲種群中的垂直傳遞及對鬆毛蟲災害的持續控製作用，通過反轉錄聚閤酶鏈式反應(RT-PCR)建立瞭DsCPV的早期檢測技術。依據傢蠶質型多角體病毒(BmCPV)質型多角體蛋白的覈苷痠序列設計一對引物，從純化的DsCPV\,BmCPV和舞毒蛾質型多角體病毒(LdCPV)的基因組dsRNA可成功地擴增齣長614bp的目的片段，從提取的健康鬆毛蟲幼蟲腸組織的DNA、舞毒蛾覈型多角體病毒(LdNPV)基因組覈痠、以及棉鈴蟲質型多角體病毒(HaCPV)基因組覈痠，未能擴增齣目的片段。DsCPV基因組dsRNA擴增片段的序列與BmCPV相應基因序列具有87%的同源性，檢測敏感度為1pg的DsCPV基因組dsRNA。由于鬆毛蟲與傢蠶、舞毒蛾相互之間食性不同，而且BmCPV和LdCPV對鬆毛蟲無感染性，即在鬆毛蟲體內不會有BmCPV病毒和LdCPV病毒的感染，因此該結果一方麵從分子水平上證實瞭DsCPV與BmCPV、LdCPV存在基因同源性，同時也錶明瞭依據BmCPV的基因序列設計引物對DsCPV基因組覈痠建立的RT-PCR擴增體繫，可以作為鬆毛蟲種群中DsCPV的一種敏感、特異、早期、快速的檢測手段。
위연구DsCPV재송모충충군중적수직전체급대송모충재해적지속공제작용，통과반전록취합매련식반응(RT-PCR)건립료DsCPV적조기검측기술。의거가잠질형다각체병독(BmCPV)질형다각체단백적핵감산서렬설계일대인물，종순화적DsCPV\,BmCPV화무독아질형다각체병독(LdCPV)적기인조dsRNA가성공지확증출장614bp적목적편단，종제취적건강송모충유충장조직적DNA、무독아핵형다각체병독(LdNPV)기인조핵산、이급면령충질형다각체병독(HaCPV)기인조핵산，미능확증출목적편단。DsCPV기인조dsRNA확증편단적서렬여BmCPV상응기인서렬구유87%적동원성，검측민감도위1pg적DsCPV기인조dsRNA。유우송모충여가잠、무독아상호지간식성불동，이차BmCPV화LdCPV대송모충무감염성，즉재송모충체내불회유BmCPV병독화LdCPV병독적감염，인차해결과일방면종분자수평상증실료DsCPV여BmCPV、LdCPV존재기인동원성，동시야표명료의거BmCPV적기인서렬설계인물대DsCPV기인조핵산건립적RT-PCR확증체계，가이작위송모충충군중DsCPV적일충민감、특이、조기、쾌속적검측수단。
In order to understand vertical transmission of DsCPV and its role in sustained controlling pine caterpillars,this study was undertaken to develop the assay technique for detection of DsCPV in early stage of infection of pine caterpillars,based on the reverse transcription of RNA followed by the polymerase chain reaction amplification (RT\|PCR).A pair of primers producing 614bp amplification fragment were designed based on the sequence of the C\|polyhedrin gene of Bombyx mori CPV (BmCPV),and the expected amplification products were obatined when genomic dsRNAs isolated from purified BmCPV,DsCPV and Lymantria dispar CPV (LdCPV) were used as templates.Genomic dsRNAs isolated from Heliothis armigera CPV and DNAs from Lymantria dispar nuclear polyhedrosis virus (LdNPV) and DNAs from midgut tissue of healthy Dendrolimus spectabilis (Walker) larva did not yield any amplification products.The detection limit of purified DsCPV genomic dsRNAs was 1.0pg.The nucleotide sequence of the 614bp DNA amplification product from DsCPV was 87% homogenous with the corresponding part of C\|polyhedrin gene of BmCPV.These results demonstrated that BmCPV,DsCPV and LdCPV were homologous as classfied in the same electropheres type I,and that this RT\|PCR assay could be used for early,rapid,sensitive and specific detection of DsPCV infection in the natural population of the pine moth,due to the apparent different feeding habits of Bombyx mori,Dendrolimus ssp.and Lymantria dispar larva each other and the no infection of BmCPV and LdCPV to Dendrolimus ssp.
