Expression of nitrite reductase gene from Lactobacillus plantarum from Sichuan pao cai

Objective: To clone and the nitrite reductase gene( nir) from Lactobacillus plantarum and detect the enzyme activity. Methods: The primers were designed according to nir sequence in Gen Bank. The DNA from Lactobacillus plantarum which came from Sichuan pao cai was extracted. The nir gene was amplified by PCR,it was cloned into T vector p MD19.The recombinant plasmid was constructed for analyzing the sequence homology.The plasmid by double enzyme digestion was connected to the expression plasmid p ET- 32a( +). It was constructed the recombinant plasmid p ET- 32a- nir. The plamid was transformed into E.coli BL( DE3) in order to express the research. The new host bacteria grew with IPTG induction,the cell was broken with ultrasonic. The crude enzyme was extracted and the enzyme activity was measured. Result: The purpose of cloned gene length was 1638 bp.A kind of recombinant protein with the molecular weight of 80 ku was obtained. The enzyme activity was 4.2 U / mg. Conclution: The nitrite reductase gene from Lactobacillus plantarum was successfully cloned and transformed in E.coli BL( DE3),the expression product had enzyme activity.