食品科学2024，Vol.45Issue(13)：89-95,7.DOI:10.7506/spkx1002-6630-20230904-025
河虾过敏原原肌球蛋白的基因克隆与原核表达
Gene Cloning and Prokaryotic Expression of the Allergen Tropomyosin from Macrobrachium nipponense
摘要
Abstract
To explore substitutes for natural tropomyosin as basic materials for the diagnosis and treatment of shrimp allergy,total RNA was extracted from Macrobrachium nipponense,and the full-length sequence of the tropomyosin gene was cloned by rapid amplification of cDNA ends(RACE).Specific primers were designed based on the cDNA sequence of tropomyosin and the expression plasmid pET-30a-TM was constructed.Finally,recombinant tropomyosin was expressed under the induction of isopropyl-β-D-thiogalactopyranoside(IPTG).The results showed that the full-length cDNA sequence of tropomyosin was 1 658 bp in length(GenBank accession number:OP974621).Its open reading frame(ORF)was 855 bp in length,encoding 284 amino acids,with a predicted protein isoelectric point of 4.70 and a predicted molecular mass of 32.8 kDa.The highest expression was obtained after 4 h of induction with a final IPTG concentration of 1 mmol/L at 37 ℃.The recombinant tropomyosin mainly existed as a soluble form in the supernatant of cell lysate with a molecular mass of approximately 38 kDa.关键词
河虾/原肌球蛋白/基因克隆/原核表达/重组蛋白Key words
Macrobrachium nipponense/tropomyosin/gene cloning/prokaryotic expression/recombinant protein分类
生物科学引用本文复制引用
骆叶晴,郑双艳,孙耀斌,陈娇,刘鑫,陈红兵,谢彦海..河虾过敏原原肌球蛋白的基因克隆与原核表达[J].食品科学,2024,45(13):89-95,7.基金项目
国家自然科学基金地区科学基金项目(32060584) （32060584）
江西省自然科学基金项目(20232BAB205084) （20232BAB205084）
