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邻苯二酚-2,3-双加氧酶基因在芘降解菌基因组的整合与表达
赵亚光, 段魏魏, 徐苗, 吴盼云, 肖璐梅, 马腾飞, 晁群芳
0
(新疆大学生命科学与技术学院)
摘要:
为了构建遗传稳定且高效降解多环芳烃工程菌,本研究通过PCR技术克隆Pseudomonas songnenensis wp3-1的邻苯二酚-2,3-双加氧酶(C23O)基因,连接于自杀性载体pUTmini-Tn5得到重组载体pUTmini-Tn5-C23O,通过三亲接合作用经mini-Tn5转座将重组载体pUTmini-Tn5-C23O中的C23O基因整合到菌株Pseudomonas sp. wp4的染色体DNA中,得到基因工程菌wp4-C23O。在不同pH值、温度下,菌株wp4和工程菌wp4-C23O对芘浓度50 mg/L进行降解7 d,结果表明:两株菌降解最适温度为37℃、最适pH值为7.5。在此条件下,工程菌wp4-C23O对芘降解率显著高于wp4菌株(p<0.05),降解率提高11.45%。
关键词:  多环芳烃(PAHs)  邻苯二酚-2,3-双加氧酶(C23O)基因  三亲接合转化  基因工程菌
DOI:
投稿时间:2018-07-17修订日期:2018-11-20
基金项目:国家自然科学基金(31460027)
Integration and Expression of Catechol-2,3-Dioxygenase gene in Pyrene degrading bacteria genome
Yaguang Zhao, Weiwei Duan, Miao Xu, Panyun Wu, Lumei Xiao, Tengfei Ma, Qunfang Chao
(College of Life Science and Technology,Xinjiang University)
Abstract:
In order to construct genetic stability and high degrading polycyclic aromatic hydrocarbons genetically engineering bacteria,In this study, the catechol-2,3-dioxygenase (C23O) gene of Pseudomonas songnenensis wp3-1 was cloned by PCR and Linked to the suicide vector pUTmini-Tn5 to obtain the recombinant vector pUTmini-Tn5-C23O,which was transformed into Pseudomonas sp. wp4 by triparental conjugatio.Finally ,C23O gene was integrated into the chromosomal DNA of wp4 and obtained genetically engineering bacteria wp4-C23O . Under different pH and temperature, strain wp4 and strain wp4-C23O were used to degrade pyrene concentration 50 mg/L to 7 d,the results showed that the optimum degradation temperature and the optimal pH value of the two strains of bacteria was 37℃、7.5,respectively.Under this condition, the engineering strain wp4-C23O for pyrene degradation rate was significantly higher than that of wp4 strain(p<0.05), the degradation rate increased by 11.45%.
Key words:  polycyclic aromatic hydrocarbons(PAHs)  catechol-2,3-dioxygenase (C23O) gene  triparetal conjugation  genetically engineering bacteria