Gene Cloning And Expression Of Helicobacter Pylori CagA Proteidic Peptide

Objective: In order to make the kit to diagnose the infection of He I icobacter pylor i with high toxigenicity,the Helicobacter pylori CagA proteidic peptide was expressed in E.coli Top 10 with expression vector ,and the most antigenic protein fragment was determined.Methods: According to the Helicobacter pylori cagA gene.five pairs of primers are designed with the DNAMAN software.Five cagA gene fragments have been amplified by PCR and inserted into the expression vector pGEX-3X.The five reformed plasmids are transformed into E.coli Top 10 by calcium-chloride-mediated transformation. By the means of PCR, endonucleases and gene-sequence,the positive plasmids are selected.The selected plasmids are induced to express the CagA proteidic peptides about 66KD, 29KD, 36KD, 99KD and 63KD using Immol/LIPTG .The fusion proteins are primarily purified by 8mol/L urea.ELISA is used to determine which proteidic peptide is the most antigenic.Results: The CagA proteidic peptides were expressed in E.coli Top 10,and the expressed proteidic peptides have antigenity.Among these peptides,66KD fragment is the most antigenic.Conclusion: The 66KD CagA proteidic peptide is the most antigenic.