Kunming Hcv Molecular Epidemiology And Core Protein Expression Plasmid

Hepatitis C virus is prevalent in worldwide. The estimated infection rate and infected population is 3% and 170 million respectively. In China, HCV infection rate is estimated to be 2.5～4.9%, and the total infected population is up to 50 million. HCV is becoming the major cause of hepatitis liver, cirrhosis and hepatocellular carcinoma. Because of the lack of effective medicine and vaccine, hepatitis C has become the second dangerous pathogen following human immunodeficiency virus (HIV).During the replication of HCV, the nucleotide substitution is common because of RNA-dependent RNA polymerase (RdRp) inducing random nucleotide errors. Based on the phylogenetic relationship, HCV could be divided into 6 major genotypes, 70 subtypes and numerous quasispecies. The distribution of HCV genotypes appeared to be distinct regional discrepancy. The clinical symptoms, disease progression and the effect of chemotherapy are related to genotypes of HCV. The research of HCV genotype is able to clarify the risk factors, the transmission of HCV, virus mutation and its evolutional characteristic. Meanwhile, HCV genotypes would offer the epidemiology evidences for vaccine and drug development, treatment adoption.Through sequencing and analysis on HCV 5'NCR-C region of 42 specimens from general population and 14 specimens from IDUs in Kunming, we found:1. The genotypes of all HCV specimens could be distinguished by this established method, including discrimination between HCV genotype 1 and 6.2. Among general population of Kunming, HCV genotype 1b and 3 were the most predominant. Genotype 2a was proved to be major genotypes. In addition, genotypes 6k and 6n were also found. This kind epidemic was different with that in other inner provinces of China (1b/2a).3. Among HIV/HCV co-infected Injection Drug Users (IDUs), HCV genotype lb and 3 were the predominant genotype. Genotype 2a was not found. It was similar with neighbored contries of southeast Asia.4. It is illuminated that specimens of genotype 1b could be devided into five groups. All the six specimens of genotype 1b from IDUs are classified into the second group. It means that HCV could be transmitted by several ways into general population. However, there is only one transmission route, HCV transmit into IDUs by it.5. There is the distinct difference of A2 restricted CTL epitopes Core protein between genotype 1b and 3b of at the region of HCV. It maybe the reason of 1b strains infected patients' fast disease progression into chronic hepatitis, hepatocirrhosis and HCC.6. The course of HCV was not related to ALT and ASL. ALT and AST were not discrepancy in different HCVgenotypes. However, HCV RNA load of genotype lb is higher than that of other genotypes. Comparing with ALT and AST, HCV RNA load is regarded as the criterion to ditermine the course of the disease.Earlier detection is very critical to control HCV prevalence. One of the most important methods is to determine the existence HCV Core protein during "window period". It is the crucial to construct Core protein expressed plasmid. In our study, HCV Core protein was cloned into pET 100/D-TOPO Vector and expressed in BL21 Chemically Competent E.coli . Then, molecular weight of protein was detected by SDS-PAGE. The anticipated protein is acquired through finding of 23KD stripe. The best activate timein this produce is is 2.5 hour.In summary, HCV molecular epidemiology in Kunming has been studied. Those results are able to clarify HCV molecular epidemiologic charasteritic, trace the origin of HCV transmittion, analysis HCV evolutional characteristic. Meanwhile, it offers the basis for clinic diagnosis, chemotherapy, drug and regional vaccine development. Expression of HCV Core protein would offer the basis for the development of HCV detection Kits.