Prokaryotic expression,purification and polyclonal antiserum preparation of the transcription factor VpSBP3 gene from Vitis pseudoreticulata

【Objective】 The objective of this study is to obtain polyclonal antiserum of the transcription factor gene VpSBP3 from Vitis pseudoreticulata for the sake of supporting the study of VpSBP3 function.【Method】 The recombinant prokaryotic expression vector pGEX-VpSBP3 of the VpSBP3 gene was constructed and translated into cells of E.coli Rosetta(DE3).The induced fusion protein with IPTG was purified by the electrodialysis,and the rabbit was immunized to obtain the polyclonal antiserum.【Result】 The recombinant vectors pGEX-VpSBP3 was constructed in E.coli DE3 and the 68 kD fusion protein of GSTVpSBP3 was efficiently expressed.Then the polyclonal antiserum was obtained by immunizing the rabbit with the purified protein.The result of Western blot indicated that the polyclonal antiserum had a specific antigen-antibody recognition feature.【Conclusion】 The results suggested that the fusion protein GST-VpSBP3 was expressed at high levels overnight under the induction condition of 0.7 mmol·L-1IPTG,27℃.The polyclonal antiserum obtained by immunizing a rabbit with the purified fusion protein had high specificity and titer,and could be used for the functional analysis of VpSBP3 gene.