Cloning,Expression and Sequence Analysis of Mn-SOD Gene of Monascus

In order to obtain the engineering bacteria and expression products of Mn-SOD in Monascus,the target gene fragment was obtained by designing the degeneracy primers of Monascus H4000,and then the full-length primers were designed to amplify the full-length sequence of Mn-SOD gene of Monascus by PCR. The target gene was digested by EcoR Ⅰ and Hind Ⅲ,and then linked to pET28a which was digested by the same enzyme,and then transformed into E. coli BL21 to induce expression. The obtained gene was predicted to encode 152 amino acids with a predicted relative molecular weight of 17 kDa. The cloned gene of Mn-SOD was compared with the NCBI database,it was found that the similarity of the sequence was 99% with that of sod gene of Monascus orange,and the sequence had higher similarity with the Mn-SOD gene of Anthracis,Aspergillus oryzae and the Aspergillus flavus. The molecular weight of Mn-SOD was estimated to be 19 kDa by SDS-PAGE. The expressed protein showed good acid resistance,which still had enzyme activity at 1 h after pH2.0 treatment.