对猪繁殖与呼吸综合征病毒易感的Marc-145细胞克隆株的筛选  

Screening of Marc-145 cell clones susceptible to porcine reproductive and respiratory syndrome virus

作　　者：谢佳芮 寇美玲 陈培富 苗海生[2] XIE Jiarui;KOU Meiling;CHEN Peifu;MIAO Haisheng(College of Veterinary Medicine,Yunnan Agricultural University,Kunming 650201,China;Yunnan Animal Science and Veterinary Institute/Yunnan Province Key Laboratory of Tropical and Subtropical Animal Virology/Key Open Laboratory of Tropical and Subtropical Animal Virology,Ministry of Agriculture,Kunming 650224,China)

机构地区：[1]云南农业大学动物医学院,云南昆明650201 [2]云南省畜牧兽医科学院/云南省热带亚热带动物病毒病重点实验室/农业部热带亚热带动物病毒学重点开放实验室,云南昆明650224

出　　处：《畜牧与兽医》2020年第11期67-71,共5页Animal Husbandry & Veterinary Medicine

基　　金：云南省技术创新人才培养对象(2018HB076)。

摘　　要：为了提高猪繁殖与呼吸综合征病毒(porcine reproductive and respiratory syndrome virus,PRRSV)在Marc-145细胞中的增殖效价,本研究旨在筛选出一株对该病毒易感度高的Marc-145细胞株。采用有限稀释法从母代细胞中分离单细胞克隆株,通过RT-PCR的方法测定毒株在克隆细胞上的核酸量,通过TCID50检测毒株对克隆细胞株的易感性,同时进行细胞生长曲线绘制。结果表明,获得2-D4、2-E6、2-C8、2-G9、2-H11株共5株Marc-145克隆细胞。接毒72 h后,2-D4、2-E6、2-C8和2-H11株细胞出现明显细胞病变,2-G9株和母代细胞培养上清液核酸检测呈阳性,但未出现细胞病变。同时,2-D4株对PRRSV毒株高度易感,该克隆细胞的分裂增殖能力要强于其他克隆株。综合上述试验结果,2-D4株更适宜于PRRSV培养,因此采用有限稀释法筛选对PRRSV易感的Marc-145细胞是一种可行的科学研究方法。This study was to improve the titers of porcine reproductive and respiratory syndrome virus(PRRSV),so as to screen a Marc-145 cell line with high susceptibility to PRRSV.Single cell clones of Marc-145 cells were isolated from the original cells by limited dilution.The quantity of the virus on cloned cells was determined by RT-PCR.The susceptibility of the strain to the cloned cell line was detected by TCID50,and the cell growth curve was drawn as well.The results showed that 5 Marc-145 cloned cells were obtained and named 2-D4,2-E6,2-C8,2-G9 and 2-H11.The cells then receive a 72 hours of virus infection,of which 2-D4,2-E6,2-C8 and 2-H11 showed cytopathic effect(CPE),while 2-G9 and the original cell culture supernatant were nucleic acid-positive,but no CPE appeared.At the same time,2-D4 was highly susceptible to PRRSV,and the proliferation of the 2-D4 cells was better than that of the other clones.In conclusion,the 2-D4 strain was more suitable for the cultivation of PRRSV,which indicated that screening Marc-145 cells susceptible to PRRSV by limited dilution in this study was a feasible research method.

关 键 词：单细胞克隆 MARC-145细胞 有限稀释法 PRRSV 易感株 

分 类 号：S852.65[农业科学—基础兽医学]