兽类学报2012，Vol.32Issue(2)：149-155,7.
犬瘟热病毒小熊猫株反向遗传系统的构建及其鉴定
Rescue of lesser panda canine distemper virus by reverse genetic manipulation
摘要
Abstract
In this study, the CDV originated from lesser pandas was isolated and served as the template for the infectious cDNA clone of CDV. After genome sequencing, seven cDNA fragments containing full-length CDV genome sequence were obtained by RT-PCR. The seven different fragments were digested and spliced together, and then inserted into eukaryotic expression vector pCI. Thus, we obtained the full-length cDNA of CDV lesser panda named pCI-CDV-LP. Furthermore, three helper plasmids were constructed by cloning the N, P, L protein ORF of lesser panda strain CDV in pCI vectors, respectively. The sequences of nuclease and CDV cDNA in pCI vector were verified by restriction enzyme digestion and sequencing. The full-length plasmid and three helper plasmids were then co-transfected into BSR cells using transfection reagent Lipofectamine (TM)2000. The results of RT-PCR, indirect immunofluorescence assay and viral infection assay showed that the lesser panda CDV was rescued successfully, and the reverse genetics of lesser panda CDV was built successfully. The results of the current investigation provide an important insight into the pathogenesis of canine distemper virus.关键词
犬瘟热病毒小熊猫株/反向遗传系统/构建Key words
Construction/ Lesser panda canine distemper virus/ Reverse genetic manipulation分类
生物科学引用本文复制引用
王磊,赵永坤,杨松涛,黄耕,高玉伟,夏咸柱,李天松,刘玉秀,冯娜,王照,姜雪,王胜乐,许微微,王铁成..犬瘟热病毒小熊猫株反向遗传系统的构建及其鉴定[J].兽类学报,2012,32(2):149-155,7.基金项目
国家科技支撑计划—动物源性人兽共患病病原与分子流行病学研究(2010BAD04B01) （2010BAD04B01）
