稳定表达犬瘟热病毒细胞受体SLAM的BHK-21/SLAM细胞系的建立  

Establishment of BHK-21 cell line stably expressing caine distemper virus recepter of signaling lymphocyte activation molecule

作　　者：朱颖[1] 刘文鑫[1] 赵姝静[1] 李丹丹[1] 冯瑜菲[1] 关玮琨[1] 何丽丽[1] 江馗语 师东方[1] 

机构地区：[1]东北农业大学动物医学学院,黑龙江哈尔滨150030 [2]沈阳市动物疫病防控中心,辽宁沈阳110161

出　　处：《中国预防兽医学报》2014年第6期440-443,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家科技支撑计划(2012BAD12B03;2012BAD12B05)

摘　　要：为建立稳定表达犬瘟热病毒(CDV)受体,即信号淋巴细胞激活因子(SLAM)的BHK-21细胞系,本研究将重组表达质粒pDisplay-SLAM转染于BHK-21细胞,通过G418压力筛选并结合有限稀释法、免疫荧光、RT-PCR和western blot筛选出稳定表达SLAM的阳性细胞(BHK-21/SLAM)。病毒感染试验结果显示,CDV在BHK-21细胞中无细胞病变(CPE)产生,而感染BHK-21/SLAM细胞12 h后即可出现CPE;BHK-21/SLAM细胞培养物上清液中的CDV核酸含量于24 h达到最高值(107.9拷贝/μL),明显高于对照组BHK-21细胞培养物上清液中的核酸含量(106.28拷贝/μL)。研究结果表明,与BHK-21细胞相比较,CDV在BHK-21/SLAM细胞中的病毒核酸含量更高,并能够产生明显的CPE。该细胞系的建立在CDV的分离、生物学特性研究和疫苗毒生产等方面具有良好应用前景。To establish a cell line stably expressing canine distemper virus （CDV） receptor of signaling lymphocyte activation molecule （SLAM）,the recombinant plasmid pDisplay-SLAM containing the SLAM gene was transfected into BHK-21 cells, and the SLAM gene integrated BHK-21（designated BHK-21/SLAM） cells were screened in present of G418, cloned by end point dilution, and identified by immunofluorescence assay, RT-PCR and western blot. The results of virus propagation showed that the CDV was able to produce CPE in BHK-21/SLAM cells, but not in the BHK-21 cells. In addition, the CDV titer in the culture medium was significantly higher in BHK-21/SLAM cells （107.9 copies/μL） than in BHK-21 cells （106.28 copies/μL） at 24 hours after inoculation. The results of this study showed that the established BHK-21/SLAM cell line could be potentially used in virus isolation, biological characteristics study and vaccine production of CDV.

关 键 词：犬瘟热病毒 信号淋巴细胞激活因子 BHK-21细胞系 BHK-21 SLAM细胞系 

分 类 号：S852.65[农业科学—基础兽医学]