1.Yeast surface display technology and its applications
ShuaiZheng JIA ; Hongyan, SUN ; Quanli, WANG
Bulletin of The Academy of Military Medical Sciences 2001;25(2):140-142
Yeast surface display is a new technology developed in recent years,which displays protein of interest on the surface of yeast cell.It can be used to display complex eukaryotic proteins requiring post-translational processing including glycosylation and efficient folding for functional activity. The basis of this technology, its development,methods of screening,its applications and perspective are mainly described in this paper.
2.Establishment of reporter gene labeled HCV full-length genome replication cell model
Bo GAO ; Shuaizheng JIA ; Jianchun PENG ; Yi WANG ; Wei FAN ; Yintai LI ; Linsheng ZHAN ; Jinbo XU
Chinese Journal of Microbiology and Immunology 2011;31(6):523-527
Objective To establish a stable HCV full-length genome replication cell model which is labeled with reporter gene and easyly to quantify intracellular HCV proteins and RNA level. Methodsneo gene was inserted into Luc-JC1 to make Luc-JC construct. Luc-JC RNA was obtained by in vitro transcription and then delivered into Huh7 cells by transfection. G418-resistant clones of Huh7 cells were obtained by selection. Clones of HCV full-length genome replication cell were confirmed by luciferase activity assay, Western blot and cleaveage of eYFP-MAVS by HCV NS3/4A protease. Then, HCV replication cell colonies were treated by different dose IFN-α in order to observe the change of luciferase activity, HCV protein and RNA level. Results At 3-4 weeks post-transfection, visible colonies were selected and stained by crystal violet. Luciferase activity and HCV NS3, NS5A protein were detected by luciferase activity assay and Western blot, respectively. Subcellular localization of eYFP-MAVS transferred from mitochondria to cytoplasms by cleavage of NS3/4A protease in cell colonies. Luciferase activity, HCV protein and RNA diminished obviously after IFN-α treatment. Conclusion A stable HCV full-length genome replication cell model labeled by reporter gene was successfully established and reporter activity can be used to indicate level of HCV proteins and RNA in cells. This cell model is a useful tool for the study on HCV pathogenesis and the screening of antiviral drugs.
3.Cloning and sequencing of partial gene from SENV isolate in China
LinSheng ZHAN ; Qinghua, MENG ; ShuaiZheng JIA ; Hongyan, SUN ; ZhiYan, DU ; Quanli, WANG
Bulletin of The Academy of Military Medical Sciences 2001;25(2):88-90
Objective:To investigate the presence of SENV infection among patients in China,and analyze partial nucleotide sequence of SENV isolated from a patient with non A-G hepatitis.Methods:A nested polymerase chain reaction (PCR) assay with primers from ORF1 of SENV genome was established to detect SENV DNA.The PCR product was cloned and sequenced.Results:SENV DNA was positive in 2 of 7 patients with non A-G hepatitis and TTV negative.Partial gene of a SENV isolate was compared with the corresponding region of SENV isolate(AX025730)from Italy and was found that the nucleotide homology was 90%.Conclusions:The results of this study confirmed the presence of SENV infection in China.The development of a PCR assay for SENV DNA detection and the cloning,sequencing of the SENV isolate have important implication for the diagnosis and epidemiological investigation on SENV infection.