Soluble expression and charaterization of anti-carbofuran single chain Fv

To express and characterize the soluble anti-carbofuran(CBF) single chain Fv(sc Fv). The gene of CBF sc Fv using plasmid p CANTAB5E-sc Fv as template,by means of designing the primers which consists of15 amino acids as the linker peptide(Gly4Ser)3,splicing by overlap extension heavy chain and light chain fragments,was cloned into p LIP6/GN vector to construct the recombinant plasmid p LIP6/GN-sc Fv. Recombinant plasmid was identified the recombinant plasmid by restrictive enzyme digestion, PCR amplification and sequence analysis. Finally,the recombinant protein was expressed in E.coli BL21,determined by SDS-PAGE and Western blot.ELISA was used to determine the antigenicity of the recombinant protein. The results showed that recombinant plasmid p LIP6/GN-sc Fv contained the inserted fragment. The expression of sc Fv linked with alkaline phosphatase showed 78 ku and could competitively combine with CBF,the IC50 was 26.80 ng/m L. The sc Fv of CBF was successful cloned and expressed,which could be useful for corresponding immunoassay methods.