中国畜牧兽医2019，Vol.46Issue(1)：10-19,10.DOI:10.16431/j.cnki.1671-7236.2019.01.002
从江香猪IFN-β基因的序列分析及原核表达
Sequence Analysis and Prokaryotic Expression of IFN-β Gene in Congjiang Xiang Pig
摘要
Abstract
This study was aimed to investigate the molecular sequences and prokaryotic expression product characteristics of interferon-beta (IFN-β) gene coding region in Congjiang Xiang pig.Congjiang Xiang pig was selected as the research object, the total RNA was extracted from liver tissue of Congjiang Xiang pig, and the mRNA was reversely transcribed into cDNA.The encoding region of IFN-βgene was amplified by specific primers, ligated into pET-28 avector, obtained the recombinant plasmid pET28a-CJpoIFN-β, and the CDS sequence of IFN-βgene was analyzed using bioinformatics softwares.The pET28a-CJpoIFN-β was transformed into Escherichia coli BL21 (DE3) competent cells.After induction by IPTG, the product of prokaryotic expression was analyzed by SDS-PAGE and Western blotting.The results showed that the CDS sequence of IFN-βgene in Congjiang Xiang pig (CJpoIFN-β) was 561 bp, which encoded 186 amino acids;CJpoIFN-βprotein was secretory protein, and the first 21 amino acid residues was predicted to be the signal peptide.The secondary structure of CJpoIFN-βprotein was mainly alpha helix (77.42％) and random coli (17.74％) .The nucleotide sequence alignment results showed that the IFN-βgene homology was 99.5％to 100.0％between Congjing Xiang pig and others porcine, the lowest homology was 35.2％ between Congjing Xiang pig and poultry.Phylogenetic tree result showed that CJpoIFN-βwas closely related to that of Meishan pig and Bama pig, the amino acid homology was100.0％in Congjiang Xiang pig and Bama pig, Meishan pig;And the homology was 99.5％ between Congjiang Xiang pig and Guizhou Baixiang pig, there were three amino acid differences in E43 Q, K73Rand C161 R.Western blotting results showed that His-tag recombinant protein could be recognized by His monoclonal antibody, the band was about 24 ku.The results of this experiment provided a reference for further study on the biological activity of IFN-βgene and speeding up the effective utilization of the resources of Congjiang Xiang pig.关键词
从江香猪/IFN-β基因/序列分析/原核表达Key words
Congjiang Xiang pig/IFN-β gene/sequence analysis/prokaryotic expression分类
农业科技引用本文复制引用
温贵兰,张升波,李昌红,徐丽,田浪,文明,王开功,程振涛,赵德刚..从江香猪IFN-β基因的序列分析及原核表达[J].中国畜牧兽医,2019,46(1):10-19,10.基金项目
国家自然科学基金(31460668) （31460668）
贵州省科学技术基金(黔科合J字[2015]2046号) （黔科合J字[2015]2046号）
贵州大学博士基金(贵大人基合字[2013]12号) （贵大人基合字[2013]12号）
贵州省高层次创新型人才培养(黔财教[2017]号) （黔财教[2017]号）
