猪内源性反转录病毒感染HEK293细胞模型的建立

Establishment and Identification of Porcine Endogenous Retrovirus-infected HEK293 Cell Model

目的 建立猪内源性反转录病毒感染HEK2 93细胞的模型 ,验证PERV在体外对人源细胞的感染性 ,并进一步研究PERV的生物学特性。方法 将G4 18抗性的HEK2 93细胞与猪源PK 15细胞共培养 ,6周后 ,用含有6 0 0 μg mlG4 18的选择性培养基对共培养细胞进行加压筛选 ,用以除去共培养体系中的PK 15细胞 ;然后应用PCR及RT PCR的方法对所制备的感染细胞模型进行系统鉴定。结果 经 6周共培养及 3周加压筛选后 ,细胞的形态、生长速度及折光性均未见明显变化 ;PCR及RT PCR鉴定表明 ,筛选后的共培养体系中已没有PK 15细胞的存在 ;PERV特异性检测方法检测显示 ,该细胞模型的DNA中已有PERV的整合且有PERV特异性mRNA的表达。结论 本实验成功建立了PERV感染HEK2 93细胞的模型 ,证实了PERV在体外对人源细胞具有感染性 ;

Objective To obtain a model of the HEK293 cells infected by PERV and further research the biological characters of the PERV. Methods The G418-resistant HEK293 cell and PK-15 cell were co-cultured in a system, the PK-15 cell in co-culture system was eliminated using the G418 selective culture medium after 6 weeks. The infected HEK293 cells were identified using PCR and RT-PCR methods. We also performed another PCR to detect existence of the PK-15 cell in co-cultured system with the species-specific primers for pgt34 intron of porcine α-1,3-galactoyltransferase gene. Results The PERV can be integrated in HEK293 cell genome and can be transferred into mRNA of PERV. No species-specific fragment of swine is amplified in co-cultured system after eliminated PK-15cells, but the control cell of PK-15 can amplifie specific fragment. These result demonstrate that the PK-15 cells are eliminated completely in co-culture system. Conclusion A model of HEK293 cell line infected by porcine endogenous retrovirus have been established, demonstrating that the PERV can infect human's cell.