适用于流感病毒感染研究的qRT-PCR内参基因评估OACSTPCD

Objective To identify stable reference genes for a comparison of the transcription levels of target host genes under viral infection in order to provide data for studies on interactions between the host and the influenza virus.Methods Reverse transcription quantitative real-time PCR(RT-qPCR)was performed to detect the relative expression levels of six candidate reference genes,including glyceraldehyde 3-phosphate dehydrogenase(GAPDH),β-actin,18S RNA,β2-microglobulin(B2M),ubiquitin-conjugating enzyme E2D2(UBE2D2),and ribosomal protein L37A(RPL37A)in classical cell models(A549 cells and THP-1 cells)under different conditions.The stability of the reference genes was evaluated using such methods as BestKeeper,GeNorm,NormFinder,and comparative A Ct method.Results The stability of reference genes varied depending on conditions.When such experimental factors as influenza virus infection and immune activation were taken into consideration,β-actin and GAPDH were identified as the most stable reference genes in A549 cells and THP-1 cells,followed by UBE2D2 and B2M.Conclusion The optimal reference genes in A549 cells and THP-1 cells under influenza virus infection or after being treated with interferons or LPS have been identified,which is of referential value for studying the mechanisms of viral infections.