摘要
口蹄疫是由口蹄疫病毒(FMDV)引起的一种高度接触性传染病,主要侵害偶蹄动物。乳鼠常作为一种重要的实验动物模型用于FMDV的研究;整联蛋白αvβ6是FMDV的重要受体之一。为深入研究整联蛋白αvβ6在FMDV感染乳鼠中所发挥的作用,克隆了乳鼠整联蛋白αvβ6的两个亚基,并将其导入中国仓鼠卵巢细胞(Chinese hamster ovary,CHO-677)基因组中,构建了稳定表达乳鼠整联蛋白αv和β6亚基的细胞系CHO-677-mαvβ6,并分别选用两种不同血清型的野生型FMDV毒株Asia1/HN/CHA/06和O/BY/CHA/2010感染细胞系来分析细胞系对FMDV的易感性。首先通过PCR和间接免疫荧光试验证明了细胞系中整联蛋白αvβ6在基因水平成功导入,在蛋白水平成功表达。然后,通过实时荧光定量RT-PCR检测病毒RNA拷贝数,并结合TCID50试验测定了代表毒株在两个细胞上的生长曲线。结果表明,与亲本细胞CHO-677相比,细胞系CHO-677-mαvβ6对FMDV更易感,从αvβ6的功能性上进一步验证了细胞系被成功构建。
Foot-and-mouth disease（ FMD） is a highly contagious disease of domestic and wild clovenhoofed animals,and its causative agent is FMD virus（ FMDV）. The suckling mouse is an important experimental animal model for FMDV and integrin αvβ6 is an important receptor of FMDV. In order to further study the role of integrin αvβ6 in FMDV infection for suckling mice,two subunit genes of integrin αvβ6 from suckling mice were cloned and transduced the Chinese hamster ovary 677（ CHO-677） cell line to stably express both suckling mouse integrin subunits αv and β6（ CHO-677-mαvβ6）. The presence of αvβ6 in CHO-677-mαvβ6 cell line was detected by PCR and indirect immunofluorescent assay（ IFA） at the gene and protein levels,respectively. In order to analyze the susceptibility of FMDV to the cell line,the wild-type strain Asia1 / HN / CHA /06 and O / BY /CHA /2010 were used to infect the cell line. Viral RNAs were detected by real-time quantitative RT-PCR.Growth curves of the representative strains in the cell line and its parental cell line were determined by TCID50 assay. The data showed that CHO-677-mαvβ6 cell line was more susceptible to FMDV than the CHO-677 cell line,indicating that CHO-677-mαvβ6 cell line was constructed successfully.
出处
《中国生物工程杂志》
CAS
CSCD
北大核心
2015年第8期23-29,共7页
China Biotechnology
基金
国家自然基金资助项目(31302118)
