Construction and Expression of Methionine-rich and Lysine-rich Fusion Gene in Bacillus natto

作     者：Zhang Shuang Luo Chao-chao Wu Cai-xia Gao Xue-jun 

作者机构：Key Laboratory of Agricultural Biological Functional Genes Northeast Agricultural University Life Science Research Center Hebei North University 

出 版 物：《Journal of Northeast Agricultural University(English Edition)》 (东北农业大学学报（英文版）)

年 卷 期：2015年第22卷第2期

页      面：22-28页

学科分类：08[工学] 09[农学] 0901[农学-作物学] 0836[工学-生物工程] 090102[农学-作物遗传育种] 

基　　金：Supported by the Funding of High Technology Project of Ministry of Science and Technology of China(863 Project 2013AA102504-03) 

主　　题：methionine-rich gene lysine-rich gene Bacillus natto prokaryotic expression 

摘      要：Methionine and lysine are restrictive essential amino acids of livestock, they are also the most attentive indexes in the feed production to carry out the quality control and quality evaluation. Their contents in feed directly affect livestock protein synthesis. Bacillus natto has excellent probiotic properties. In this experiment, we used the genetic engineering method, fusion PCR technique, to connect methionine-rich gene （zein） from maize endosperm protein with lysine-rich gene （Cflr） from the pepper anther, then the fusion gene was inserted into the expression vector pHT43, and the recombinant plasmid pHT43/zein-Cflr was constructed. The recombinant plasmid was transferred into Bacillus natto, and induced by IPTG for the expression of the fusion gene. We found an apparent band at 40 ku site for the recombinant strain by SDS-PAGE. The contents of methionine and lysine were individually detected with HPLC, the quantities of methionine and lysine in the recombinant strain increased by 18.37% and 24.68% than the wild one, respectively. We also verified the stability of the recombinant bacterium during passaging, and found the stability was 100%. This study provided research-basis for the application of the recombined Bacillus natto as feed additive.