Development of an anhydrotetracycline-inducible gene expression system for solvent-producing Clostridium acetobutylicum: A useful tool for strain engineering

被引：46

作者：

Dong, Hongjun ^{[1
]}

Tao, Wenwen ^{[1
]}

Zhang, Yanping ^{[1
]}

Li, Yin ^{[1
]}

机构：

[1] Chinese Acad Sci, Inst Microbiol, Beijing 100101, Peoples R China

来源：

METABOLIC ENGINEERING | 2012年 / 14卷 / 01期

基金：

国家高技术研究发展计划(863计划);

关键词：

Clostridium acetobutylicum; Inducible gene expression systems; Tetracycline regulatory system; Metabolic engineering tools; COUNTER-SELECTABLE MARKER; ESCHERICHIA-COLI; TET REPRESSOR; PROTEIN; INACTIVATION; MAZF; CONSTRUCTION; MANIPULATION; PROMOTERS; THIOLASE;

D O I：

10.1016/j.ymben.2011.10.004

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

Clostridium acetobutylicum is an important solvent (acetone-butanol-ethanol) producing bacterium. However, a stringent, effective, and convenient-to-use inducible gene expression system that can be used for regulating the gene expression strength in C acetobutylicum is currently not available. Here, we report an anhydrotetracycline-inducible gene expression system for solvent-producing bacterium C acetobutylicum. This system consists of a functional chloramphenicol acetyltransferase gene promoter containing tet operators (tetO), Pthl promoter (thiolase gene promoter from C acetobutylicum) controlling TetR repressor expression cassette, and the chemical inducer anhydrotetracycline (aTc). The optimized system, designated as pGusA2-2tetO1, allows gene regulation in an inducer aTc concentration-dependent way, with an inducibility of over two orders of magnitude. The stringency of TetR repression supports the introduction of the genes encoding counterselective marker into C acetobutylicum, which can be used to increase the mutant screening efficiency. This aTc-inducible gene expression system will thus increase the genetic manipulation capability for engineering C acetobutylicum. (C) 2011 Elsevier Inc. All rights reserved.

引用

页码：59 / 67

页数：9

共 50 条

[1] Synthetic biology: new engineering rules for an emerging discipline [J].

Andrianantoandro, Ernesto ;

Basu, Subhayu ;

Karig, David K. ;

Weiss, Ron .

MOLECULAR SYSTEMS BIOLOGY, 2006, 2 (1) :2006.0028

[2] The application of Tet repressor in prokaryotic gene regulation and expression [J].

Bertram, Ralph ;

Hillen, Wolfgang .

MICROBIAL BIOTECHNOLOGY, 2008, 1 (01) :2-16

[3]

BRADFORD MM, 1976, ANAL BIOCHEM, V72, P248, DOI 10.1016/0003-2697(76)90527-3

[4] Refinement and standardization of synthetic biological parts and devices [J].

Canton, Barry ;

Labno, Anna ;

Endy, Drew .

NATURE BIOTECHNOLOGY, 2008, 26 (07) :787-793

[5] Complete nucleotide sequence of Tn10 [J].

Chalmers, R ;

Sewitz, S ;

Lipkow, K ;

Crellin, P .

JOURNAL OF BACTERIOLOGY, 2000, 182 (10) :2970-2972

[6] Tetracycline antibiotics: Mode of action, applications, molecular biology, and epidemiology of bacterial resistance [J].

Chopra, I ;

Roberts, M .

MICROBIOLOGY AND MOLECULAR BIOLOGY REVIEWS, 2001, 65 (02) :232-+

[7] One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products [J].

Datsenko, KA ;

Wanner, BL .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2000, 97 (12) :6640-6645

[8]

Desai RP, 1999, APPL ENVIRON MICROB, V65, P936

[9] Engineering Clostridium Strain to Accept Unmethylated DNA [J].

Dong, Hongjun ;

Zhang, Yanping ;

Dai, Zongjie ;

Li, Yin .

PLOS ONE, 2010, 5 (02)

[10] Controlling gene expression in mycobacteria with anhydrotetracycline and Tet repressor [J].

Ehrt, S ;

Guo, XZV ;

Hickey, CM ;

Ryou, M ;

Monteleone, M ;

Riley, LW ;

Schnappinger, D .

NUCLEIC ACIDS RESEARCH, 2005, 33 (02) :e21

← 1 2 3 4 5 →