tailieunhanh - Báo cáo y học: " A reporter system for assaying influenza virus RNP functionality based on secreted Gaussia luciferase activity"

Tuyển tập báo cáo các nghiên cứu khoa học quốc tế ngành y học dành cho các bạn tham khảo đề tài: A reporter system for assaying influenza virus RNP functionality based on secreted Gaussia luciferase activity | Zhu et al. Virology Journal 2011 8 29 http content 8 1 29 J VIROLOGY JOURNAL METHODOLOGY Open Access A reporter system for assaying influenza virus RNP functionality based on secreted Gaussia luciferase activity Wenfei Zhu1 Jianfang Zhout Kun Qin Ning Du Liqi Liu Zaijiang Yu Yun Zhu Wenhong Tian Xiaobing Wu Yuelong Shu Abstract Background Influenza A virus can infect a wide variety of animal species including humans pigs birds and other species. Viral ribonucleoprotein vRNP was involved in genome replication transcription and host adaptation. Currently firefly luciferase Fluc reporter system was used in vRNP functional assay. However its limitation for the testing by virus infection resulted in an increased need for rapid sensitive and biosafe techniques. Here an influenza A virus UTR-driven gene reporter for vRNP assay based on secreted Gaussia luciferase Gluc activity was evaluated. Results By measuring Gluc levels in supernatants reporter gene activity could be detected and quantitated after either reconstitution of influenza A virus polymerase complex or viral infection of 293T and A549 cells respectively. As compared with Fluc reporter Gluc-based reporter was heat-tolerant 65 C for 30 min and produced 50-fold higher bioluminescent activity at 24 h posttransfection. Signals generated by Gluc reporter gene could be detected as early as 6 h post-infection and accumulated with time. Testing by viral infection stronger signals were detected by Gluc reporter at a MOI of than that of 1 and the effects of PB2-627K E or amantadine on influenza vRNP activity were elucidated more effectively by the Gluc reporter system. Conclusions This approach provided a rapid sensitive and biosafe assay of influenza vRNP function particularly for the highly pathogenic avian influenza viruses. Background Bioluminescence reporters such as firefly click-beetle Renilla Gaussia Cypridina luciferases and calcium-activated photoproteins are widely used in molecular .