Cloning And Prokaryotic Expression Of Tm18 Gene Of Taenia Multiceps Oncospheres

Coenurosis,which is a kind of serious fatal zoonoses,is caused by the parasitism of Coenurus cerebralis,and develops in the central nervous system of sheep and other ungulates.Coenurosis has been documented in scattered foci throughout the world, particuly in pastureland. Drug treatment of Coenurosis is relatively conservative, the effect often unstable; surgical treatment although the prognosis is better, but only applies to the brain surface of the cysts. As people on food safety concerns and the parasite drug resistance appearance, immunization regain attention. Early vaccine antigens are natural products,can prevent the Coenurus cerebralis infection effectively; However, because the antigen originates limited, it very difficult to promote. In this condition, the genetic engineering vaccine to be a good choice.In this study,we aimed to research tick molecules for vaccine candidates.1 pair of gene special primers were designed based on the published nucleotide sequence of the 18 kDa antigens of Taenia spp.total RNA isolated from actived oncosphere, was used as template to generate complementary DNA by reverse transcription.the 552bp DNA fragment was amplified by polymerase chain reaction,and clones it into PMD18-T vector.DNA sequencing confirmed the inserted fragment was 18 kDa antigens gene, designated Tm18. The complete ORF is 396bp,encoding a 131 amino acid. By blasting the homologous sequence in GeneBank databases, the sequence of Tm18 gene of China Taenia multiceps is 91.26% percent identity to Turkey. Taenia multiceps, and the homology between deduced amino acids of Tm18 gene was 79.39%.By the technology of DNA recombination,the Tm18 gene was subcloned to the pGEX-4T-1 expression vector and highly expressed in E.coli host BL21(DE3) under 0.6 mmol/L IPTG induction 8 hours at 37℃. The expressed proteins fused by GST with the molecular mass of 39.4 kDa, was found to be sequestered into inclusion bodies. Recombination proteins expressed in Escherichia coli was recognized by sera of goats infected Coenurusce cerebralis.The recombination proteins with FAC was made into vaccine and injected mice,and anti Tml8 antibody in serum was screened by indirect ELISA.The mice immunized with recombination protein showed higher IgG,IgG1and IgG2a antibody responses, compared with the control groups.The Tm18 recombination proteins was able to inducte antibody production,and it could be a good candidate for the vaccine development.