Cloning Expression And Biological Functional Analysis Of The ADGF-B In Aedes Albopictus

Objective: We acquire r Ab-ADGF-B protein of Aedes albopictus and explore its biological function.Methods: The ORF of the Ab-ADGF-B(Gen Bank No.AAV90660.1)was downloaded from the NCBI and analyzed by bioinformatics software.Then a recombinant Ab-ADGF-B p ET32a(+)was constructed and transformed into E.coli Rosetta-gami B(DE3);the recombinant protein induced by IPTG was identified by SDS-PAGE and western blotting.After that,the antibody was prepared by immunized rabbit with purified r Ab-ADGF-B and the titer of the antibody was detected by indirect enzyme-linked immunosorbent assay(ELISA).ADA activity of r Ab-ADGF-B was measured by spectrophotometric assay and the activity of cell growth factor was detected by direct counting method of cell.The anti-coagulation of r Ab-ADGF-B was performed on human whole blood by prothrombin time(PT),thrombin time(TT),and activated partial thromboplastin time(APTT).Results: A recombinant Ab-ADGF-B-p ET32a(+)was constructed and a total of 1545 bp encoding mature peptide of Ab-ADGF-B was obtained.An ADGF domain was identified by domain prediction.A r Ab-ADGF-B of soluble protein,induced by 0.4m M IPTG at 20?,150 rpm for 24 h,was verified by western blotting and SDS-PAGE.100?mol adenosine was catalyzed to inosine by 0.67?mol r Ab-ADGF-B at 30? and p H 7.0.At the same time,the cell amount increased from5�104 to 14�105 when 0.134 ?mol r Ab-ADGF-B protein was incubated with ANA cell for 36 hours.The PT,TT and APTT were 13.33 seconds,15.23 seconds and 35 seconds respectively when 0.134 ?mol r Ab-ADGF-B protein was mixed with the human whole blood.Conclusion: The r Ab-ADGF-B protein is successfully expressed and own ADA activity and cell proliferation activity.