Abstract: To construct the prokaryotic expression plasmid of human spermidine/spermine N1-acetyltransferase (SSAT) and to induce and purify the recombined SSAT protein in E.coli JM109 (DE3). MethodsThe total RNA was extracted from colon cancer tissues and a 513bp cDNA was amplified by RT-PCR with two primers, which span the coding region of SSAT. The synthesized cDNA was subcloned into the prokaryotic expression vector pTriEx-4. The constructed vector pTriEx-4-SSAT was transformed into competence E. coli JM109 (DE3) and then was induced by IPTG. The protein was verified by SDSPAGE and Western blot with the anti HisTag monoclonal antibody. The fusion protein was purified by the Ni-NTA chromatographic column. ResultsThe prokaryotic expression plasmid pTriEx-4-SSAT was successfully constructed, and identified by digestion and sequence. An approximate 20kD exogenous protein was observed on the SDS-PAGE. The protein was verified by Western blot with an anti His-Tag monoclonal antibody. The fusion protein including 6His-Tag was purified by the NiNTA chromatographic column. ConclusionThe SSAT prokaryote expression vector was successfully constructed and the purified protein can be applied for further research of the immunity of SSAT.

Key words: Spermidine/spermine N1acetyltransferase, Prokaryote expression, Colorectal neoplasms