Preparations of prokaryotic expression system,standard protein and antiserum of human Y-box binding protein 1

Aim: To construct a GST-expression system of human Y-box binding protein 1(YB-1),prepare the YB-1 standard protein and its antiserum.Methods: The code sequence of YB-1 was subcloned to the expression vector pGEX-6P-1.The recombinant vector was transformed into E.coli BL21 to express fusion protein GST-YB1.SDS-PAGE was applied to analyze the expression level and form of fusion protein.Then,YB-1 standard protein was obtained by the means of GST-affinity chromatography and column-protease-digestion.The rabbit was immunized with YB-1 protein to prepare the anti-YB1 polyclonal antibody.Results: The recombinant expression vector was constructed.SDS-PAGE and Western blot results showed that the GST-fusion protein was high-level expressed with soluble-form,and YB-1 standard protein and its antiserum were obtained successfully.Conclusions: An economical,rapid method to prepare YB-1 standard protein is established,and further obtained the high titer and affinity YB-1 polyclonal antibody,which lay foundations for preparation of YB-1 monoclonal antibody and development of YB-1 quantitative analysis methods.