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目的:构建携带增强型绿色荧光蛋白基因(Enhanced green fluorescent protein,EGFP)的HIV-1假病毒;通过HIV-1假病毒感染未成熟树突状细胞(immatural dendritic cell,iDC)实验探讨病毒与细胞的相互作用。方法:利用慢病毒包装系统构建携带EGFP基因的HIV-1假病毒,通过RT-PCR扩增HIV-1假病毒中的EGFP基因;HIV-1假病毒感染iDC,对HIV-1假病毒感染iDC后的基因组整合和转录水平进行检测,观察被感染iDC中EGFP基因的表达。结果:构建的HIV-1假病毒通过RT-PCR结果显示扩增得到EGFP基因;HIV-1假病毒感染iDC后,在基因组整合和转录水平检测中都扩增得到了EGFP基因,荧光显微镜观察被HIV-1假病毒感染的iDC,观察到iDC表达绿色荧光蛋白。结论:成功构建携带EGFP基因的HIV-1假病毒;HIV-1假病毒可以感染iDC,并将其遗传物质整合到iDC基因组中,并经过转录和翻译使iDC表达EGFP。
OBJECTIVE: To construct HIV-1 pseudovirus carrying enhanced green fluorescent protein (EGFP) and investigate the effect of HIV-1 pseudovirus infection on immatural dendritic cells (iDC) Interaction. Methods: HIV-1 pseudoviruses carrying EGFP gene were constructed by lentiviral packaging system and EGFP gene in HIV-1 pseudovirus was amplified by RT-PCR. HIV-1 pseudovirion was used to infect iDC, After genomic integration and transcriptional level were detected to observe the expression of EGFP gene in infected iDCs. Results: The constructed HIV-1 pseudoviruses amplified the EGFP gene by RT-PCR. After HIV-1 pseudovirus infected iDC, the EGFP gene was amplified in both genomic integration and transcriptional level. Fluorescence microscopy was performed HIV-1 pseudovirion-infected iDCs, iDCs were observed to express green fluorescent protein. CONCLUSION: The HIV-1 pseudovirus carrying EGFP gene was successfully constructed. The HIV-1 pseudovirus can infect iDCs and integrate their genetic material into iDC genome. After transcription and translation, iDCs express EGFP.