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中国农学通报

中国农学通报 ›› 2015, Vol. 31 ›› Issue (14): 83-88.doi: 10.11924/j.issn.1000-6850.casb14120202

所属专题： 生物技术

• 生物技术科学 • 上一篇    下一篇

Klebisella oxytoca HD79乙酸激酶部分基因(ack)的克隆与序列分析

叶广彬1,宋 刚1,凌宏志1,葛菁萍1,平文祥1,2   

  1. (1黑龙江大学生命科学学院,微生物省高校重点实验室,哈尔滨 150080；2农业微生物技术教育部工程研究中心,哈尔滨 150500）
  • 收稿日期:2014-12-30 修回日期:2015-04-09 接受日期:2015-02-25 出版日期:2015-06-02 发布日期:2015-06-02
  • 通讯作者: 葛菁萍
  • 基金资助:
    国家自然科学基金青年科学基金项目“酸菜发酵生态系统中细菌群落与代谢物组特征耦合机制”(31300355)；黑龙江大学杰出青年基金“蜡状芽孢杆菌(Bacillus cereus) HDYM-02亚麻生物脱胶活性研究”(JCL201305)；哈尔滨市科技局青年后备人才项目“功能性乳酸菌发酵酸菜生态学过程研究”(2014RFQXJ101)；国家自然科学基金面上项目“菌群演替与温水沤麻系统关键酶产生菌代谢组学特征的耦合机制”(31270534)；黑龙江省高等学校科技创新团队“黑龙江省高等学校科技创新团队”（农业微生物发酵技术，2012td009）。

Sequence Analysis and Clone of the Partial Acetate Kinase Gene (ack) from Klebisella oxytoca HD79

Ye Guangbin1, Song Gang1, Ling Hongzhi1, Ge Jingping1, Ping Wenxiang1, 2   

  1. (1Key Laboratory of Microbiology, Life Science Department, Heilongjiang University, Harbin 150080;2Engineering Research Center of Agricultural Microbiology Technology, Ministry of Education, Heilongjiang University, Harbin 150050)
  • Received:2014-12-30 Revised:2015-04-09 Accepted:2015-02-25 Online:2015-06-02 Published:2015-06-02

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摘要： 乙酸是2,3-丁二醇典型代谢途径中的主要副产物之一，它的大量生成能够造成环境pH值的改变并且抑制菌体的生长，并对2,3-丁二醇的代谢产生影响。乙酸激酶是乙酸生成的关键酶，敲除其相关基因，理论上可以减少乃至消除乙酸的产生，从而提高2,3-丁二醇的产量和得率。本研究根据Klebisella oxytoca KCTC1686和Klebisella oxytoca HD79乙酸激酶基因ack序列的相似性设计引物，以产酸克雷伯氏菌基因组DNA和质粒载体pT-ack为模板，利用PCR克隆得到了乙酸激酶部分基因ack，长度为856 bp，无碱基缺失。ORF分析发现该部分基因没有完整的开放阅读框，但是全部位于Klebisella oxytoca KCTC1686 ack基因的开放阅读框内。结果表明该片段为Klebisella oxytoca HD79乙酸激酶部分基因序列，为后续构建产酸克雷伯氏菌乙酸激酶突变株奠定了基础。

关键词: 森林火险天气, 森林火险天气, 中短期预报, 人工神经网络, NMC指数

Abstract: Acetic acid is one of the major metabolic by-products in 2,3-butanediol classical metabolic pathway. Production of 2,3-butanediol can be affected by synthesis of numerous acetic acid which will lead to changing of environmental pH and inhibition of cell growth. Acetate kinase is a key enzyme in acetic acid metabolic pathway, knock-out of encoding genes of acetate kinase will theoretically decrease and even remove the production of acetic acid, thus the yield and productivity of 2,3-butanediol will be enhanced. In this paper, 856 bp of partial kinase gene ack without base deletion was cloned taking genomic DNA of Klebsiella oxytoca and plasmid pT-ack as templates with primers designed according to similarity of ack sequences from Klebisella oxytoca KCTC1686. The ack gene was located in open reading frame of ack gene of Klebisella oxytoca KCTC1686 though integrated ORF was not found in it by ORF analysis. The result demonstrated that the ack gene was partially acetate kinase gene from Klebisella oxytoca HD79 which laid the foundation for further construction of acetate kinase mutant of Klebisella oxytoca HD79 and provided reference for other relative researches.