ABSTRACT
<p><b>OBJECTIVE</b>To study the effect of gene optimization on the expression and purification of HDV small antigen produced by genetic engineering.</p><p><b>METHODS</b>Based on the colon preference of E. coli, the HDV small antigen original gene from GenBank was optimized. Both the original gene and the optimized gene expressed in prokaryotic cells, SDS-PAGE was made to analyze the protein expression yield and to decide which protein expression style was more proportion than the other. Furthermore, two antigens were purified by chromatography in order to compare the purity by SDS-PAGE and Image Lab software.</p><p><b>RESULTS</b>SDS-PAGE indicated that the molecular weight of target proteins from two groups were the same as we expected. Gene optimization resulted in the higher yield and it could make the product more soluble. After chromatography, the purity of target protein from optimized gene was up to 96.3%, obviously purer than that from original gene.</p><p><b>CONCLUSION</b>Gene optimization could increase the protein expression yield and solubility of genetic engineering HDV small antigen. In addition, the product from the optimized gene group was easier to be purified for diagnosis usage.</p>
Subject(s)
Electrophoresis, Polyacrylamide Gel , Genetic Engineering , Methods , Hepatitis D , Diagnosis , Hepatitis delta Antigens , Genetics , Recombinant ProteinsABSTRACT
<p><b>OBJECTIVE</b>To investigate the seroprevalence of hepatitis D virus in Foshan of Guangdong province, to provide the data for the study about it in China.</p><p><b>METHODS</b>ELISA kits from two different companies were used for detecting anti-HDV IgG of all the serum samples, and then RT-PCR was carried out about the selected serum to ensure the results. All the serum samples were collected in 2011 in The First People's Hospital of Foshan.</p><p><b>RESULTS</b>The results from two ELISA kits and RT-PCR were identical. Eight samples were positive.</p><p><b>CONCLUSIONS</b>The seroprevalence rate of HDV in Foshan is higher than that in China. It has no statistically significant difference between female and male. Morever, the older with HBsAg are susceptible to HDV.</p>
Subject(s)
Adolescent , Adult , Aged , Child , Child, Preschool , Female , Humans , Male , Middle Aged , China , Epidemiology , Coinfection , Epidemiology , Enzyme-Linked Immunosorbent Assay , Hepatitis B , Epidemiology , Hepatitis B Surface Antigens , Blood , Hepatitis D , Epidemiology , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
<p><b>OBJECTIVE</b>To establish a novel improved loop-mediated isothermal amplification (LAMP) technique to detect hepatitis A virus (HAV).</p><p><b>METHODS</b>A novel improved LAMP assay based on the addition of an acceleration primer was developed for hepatitis A virus nucleotide acid detection.</p><p><b>RESULTS</b>Precision and reproducibility analysis proved its high stability and reliability. Comparison between the improved and conventional LAMP assays revealed that the former was more sensitive with a detection limit of 5 TCID50/ml. The novel detection method displayed 100% consistency with the TaqMan real-time PCR assay when applied to clinical specimens collected from patients with confirmed acute HAV infection.</p><p><b>CONCLUSION</b>This novel technique is widely applicable as a simple diagnostic tool in the clinical field as well as for the surveillance and investigation of the infectious disease in developing countries where HAV is endemic.</p>
Subject(s)
Humans , DNA Primers , Genetics , Hepatitis A , Virology , Hepatitis A virus , Genetics , Nucleic Acid Amplification Techniques , Methods , RNA, Viral , Genetics , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To prepare HDAg with biological activities as a candidate of diagnostic reagent.</p><p><b>METHODS</b>To synthesize HDV gene fragment after codon optimization. To construct a thio-fused recombinant plasmid based on M48 expression vector. To express in E. coli induced by IPTG. To purify the protein by affinity chromatography followed by characterization in ELISA:</p><p><b>RESULTS</b>Plasmid construction was verified by enzyme digestion. SDS-PAGE indicated the molecular weight of the protein was the same as we expectation. ELISA proved its affinity with HDV antibodies.</p><p><b>CONCLUSION</b>HDAg was obtained successfully and it will pave the road to the research of HDV diagnostic reagent.</p>