HCV Loop-mediated Isothermal Amplification Detection Methods To Establish And Codon Usage Characteristic Analysis

27HCV International reference strain complete genomic sequences representing seven genotypes reported in GeneBank were blasted to designate the subject sequence of HCV RT-LAMP. according to the conserved sequence of Core protein and5-UTR of HCV(GV451224), primers FIP, BIP, F3and B3were designed, and the usefulness of reverse transcription loop-mediated isothermal amplification (RT-LAMP) for the rapid diagnosis of hepatitis C virus (HCV) RNA was evaluated. This assay showed higher sensitivities than that of nested RT-PCR, with a detection limit of600IUmL-1, and no cross-reactivity was observed with hepatitis A virus, hepatitis B virus and hepatitis E virus. Furthermore,106stored sera from recently diagnosed cases were retrospectively investigated with real-time RT-PCR, the nested RT-PCR, in parallel with this new assay. The general detection rates of HCV RT-LAMP, real-time PCR and the nested RT-PCR for106stored sera samples were95%,96%and88%, respectively. This study provides the first data on the usefulness of HCV RT-LAMP in the diagnosis of HCV RNA, especially in the early clinical diagnosis of acute HCV infection.The relative synonymous codon usage (RSCU) values, aromaticity and hydrophobicity of each polyprotein of the virus, effective number of codons (ENC) values and nucleotide contents of144HCV complete genomic sequences were calculated to implement a comparative analysis to evaluate the dynamics of the virus evolution by Codon W and SPSS. The RSCU values of each codon of144HCV ORFs indicated that all abundant codons were C/G-ended codons. The plots of principal component analysis based on sub-genotype of HCV indicated that sub-genotype1a and1b separated clearly on the axis of f2suggesting that the codon usage bias between sub-genotype1a and1b strains was different. By comparing the codon usage between HCV and human cells, we found that the synonymous codon usage pattern of HCV was a mixture of coincidence and antagonism to that of host cells. The characteristics of the synonymous codon usage patterns and nucleotide contents of HCV, and the correlation analysis between GC3s,GCl,2s, GC%(ORF), GC%(50-UTR), GC%(30-UTR), aromaticity, hydrophobicity and ENC value, respectively, indicated that mutational pressure was the dominant factor accounting for the codon usage variation and selection pressure also accounted for HCV codon usage pattern.