论文部分内容阅读
目的探讨用双嗜性逆转录病毒载体将外源基因导入日本血吸虫(Sj)细胞的生物学理论与实验依据。方法用生物信息学方法对双嗜性逆转录病毒rRam-1受体同源性分布、结构与功能作系统的分析与比较;利用携带外源E77.43基因的双嗜性逆转录病毒感染Sj童虫培养细胞,经PCR和RT-PCR检测感染细胞目的基因(E77.43)的整合与表达。结果根据生物信息学分析结果推断,Sj细胞膜上存在的SjCHGC09605和SjCHGC05362两种蛋白为非分泌性跨膜蛋白,可能具有细胞膜离子转运通道或受体蛋白的功能及双嗜性逆转录病毒感染的膜受体样作用,可能参与病毒对细胞的吸附和穿入过程;利用携带外源E77.43基因的双嗜性逆转录病毒感染Sj童虫培养细胞后,用PCR及RT-PCR检测到目的基因整合与表达,扩增的目的片段大小为330 bp,与理论值相符。结论用载有E77.43基因的双嗜性逆转录病毒感染Sj童虫细胞获得成功,推测SjCHGC09605和SjCHGC05362两种与rRam-1受体同源的蛋白可能是Sj感染过程中起作用的分子。研究结果为下一步用双嗜性逆转录病毒载体转导永生化基因到Sj细胞提供了生物学理论与实验依据。
Objective To investigate the biological theory and experimental basis of introducing exogenous genes into Schistosoma japonicum (Sj) cells with an ampicillin retroviral vector. METHODS: The homology distribution, structure and function of rMam-1 receptor were analyzed and compared systematically by bioinformatics methods. Sj (superscript +) was infected by the amphotropic retrovirus carrying exogenous E77.43 gene Schistosoma japonicum cells were cultured, and the integration and expression of the target gene (E77.43) in infected cells were detected by PCR and RT-PCR. Results According to the results of bioinformatics analysis, it was inferred that the two proteins SjCHGC09605 and SjCHGC05362 existing on Sj cell membrane were non-secreted transmembrane proteins, which may have the function of membrane ion transport channel or receptor protein and the membrane of amphotropic retrovirus infection Receptor-like role, may be involved in the virus on the cell adsorption and penetration process; using exogenous E77.43 gene ampicillin retrovirus infected Sj squirrels cultured cells, PCR and RT-PCR detected the target gene Integration and expression, amplification of the target fragment size of 330 bp, consistent with the theoretical value. Conclusion SjCHGC09605 and SjCHGC05362 are homologous to the rRam-1 receptor, suggesting that SjCHGC09605 and SjCHGC05362 may be involved in the pathogenesis of Sj infection. The results provided the biological theory and experimental basis for the next step of transducing immortalized genes into Sj cells by using an amphoteric retroviral vector.