Cloning, Expression And Preliminary Identification Of MOMP For Chlamydia Trachomatis E Serotype

Infection of human urogenital tract caused by Chlamydia trachomatis(C.t)is more and more these years.Asymptomatic and chronic C.t infection may result in severe complications such as prostatitis,infertility,ectopic pregnancy and chronic pelvic inflammatory disease etc.C.t is remarked for its state of protraction, refractoriness,easy relapse,and striking consumption.Study on C.t infection has been one of the popular topics of the world.It's urgent to improve the prevention and control measures for C.t prevalence.Research on C.t vaccine is the key point of prevention measures.Research has indicated that Chlamydia trachomatis major outer membrane protein epitope decided to produce protective antibodies,and specially as the target antigen of the antibodies.MOMP,its molecular mass about 40×10~3,is the major components of Chlamydia outer membrane complex and about more than 60 percent in the total outer membrane protein.Objective:To construct the ompl-pGEX6p-1 recombinant and induce the expression of C.t serotype E MOMP,then preliminary identification.Methods:Extracted DNA sequence of C.t E serotype which coding MOMP as a template,designing primer and amplifing the ompl genes according it.Ompl and pGEX6p-1 were linked to be recombinant plasmid ompl-pGEX6p-1, transforming into E.coli BL21.Select one colony which grows on the plate with antibiotics,cultivate it in LB medium,and extract its plasmid.Enzymes cutting and 1 %agarose gel electrophoresis can preliminary identify that recombinant plasmid is right.Induce the E.coli with the plasmid to get the target protein.By SDS-PAGE,we can recognize the target protein is in the surpernant or sediment.During western blot we use the GST-tag antibody as the first antibody and then confirm the protein can be detected.Results:The sequence of gene which got from PCR are similar with ompl of C.t E serotype in Genebank about 99.92%,showing that we have got ompl of C.t E serotype.After enzymes cutting,we got two parts of gene sequence,1200bp and 5000bp separately.We can draw a conclusion that recombinant protein MOMP of C.t E serotype was constructed successfully.The target protein induced is about 53KD and can be detected by GST-tag antibody.Sequence of recombinant plasmid showed there had a mutation in 736 basic group and the expression of protein was ended.Conclusions:(1)Ompl gene of C.t E serotype has been cloned and recombinant ompl-pGEX6p-1 has been constructed successfully;(2)The protein recombinant plasmid expressed can be detected by GST-tag antibody;(3)Sequence of recombinant plasmid showed that the taget protein has immunogenicity theoretically.