Tet-on, or Tet-off, that is the question: Advanced conditional gene expression in Aspergillus

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	Tet-on, or Tet-off, that is the question: Advanced conditional gene expression in Aspergillus
	Wanka, Franziska 1; Cairns, Timothy 1; Boecker, Simon 1; Berens, Christian 2; Happel, Anna 3; Zheng, Xiaomei 4; Sun, Jibin 4; Krappmann, Sven 3; Meyer, Vera 1
刊名	FUNGAL GENETICS AND BIOLOGY
	2016-04-01
卷号	89 页码:72-83
关键词	Aspergillus Tet-on Tet-off Conditional expression Heterologous gene expression Genetic engineering
英文摘要	In Aspergillus, controlled gene expression is often achieved using the reverse tetracycline-controlled transactivator (rtTA) dependent Tet-on system, whereby transcription is activated in a titratable manner by addition of the tetracycline derivative doxycycline. The complementary Tet-off system utilises the tetracycline-controlled transactivator (tTA) component to quantitatively reduce gene expression. In this study, we utilised a synthetic biological approach to engineer highly optimised Tet-off conditional expression systems in Aspergillus niger and Aspergillus fumigatus. Steps for delivery of these tools include utilising codon optimised cassette components, testing several promoters for improved genetic stability and validating two modified luciferase reporters for highly accurate measurements of gene expression. The Tet-off cassettes developed in this study enable facile and quantitative functional analysis, as validated by Tet-off analysis of genes involved in chitin synthesis and cell wall polarity in A. niger, and para-aminobenzoic acid synthesis in A. fumigatus. We also used a racA(G18V) dominant allele to demonstrate that Tet-off in A. niger enables gene over-expression and downregulation in a single isolate. Additionally, we used the improved luciferase reporters to show that the Tet-off cassette in A. niger enables quantification of gene oscillations. In order to demonstrate that synthetic biological approaches developed here are broadly applicable to engineering transcriptional circuits in filamentous fungi, we used our strategy for improving cassette stability by promoter replacement in the A. niger Tet-on system, which resulted in a modified Tet-on cassette with higher stability in recipient genomes. (C) 2015 Elsevier Inc. All rights reserved.
WOS标题词	Science & Technology ; Life Sciences & Biomedicine
类目[WOS]	Genetics & Heredity ; Mycology
研究领域[WOS]	Genetics & Heredity ; Mycology
关键词[WOS]	STRESS-RESPONSE ; CELL-WALL ; FUMIGATUS ; NIGER ; IDENTIFICATION ; MARKER ; RECOMBINATION ; LUCIFERASE ; STRAINS
收录类别	SCI
语种	英语
WOS记录号	WOS:000372666400009
内容类型	期刊论文
源URL	[http://124.16.173.210/handle/834782/2236]
专题	天津工业生物技术研究所_系统与合成生物技术实验室孙际宾_期刊论文
作者单位	1.Berlin Univ Technol, Inst Biotechnol, Dept Appl & Mol Microbiol, Gustav Meyer Allee 25, D-13355 Berlin, Germany 2.Friedrich Loeffler Inst, Inst Mol Pathogenesis, Jena, Germany 3.Univ Erlangen Nurnberg, Univ Klinikum Erlangen, Mikrobiol Inst Klin Mikrobiol Immunol & Hyg, Erlangen, Germany 4.Chinese Acad Sci, Tianjin Inst Ind Biotechnol, Tianjin 300308, Peoples R China
推荐引用方式 GB/T 7714	Wanka, Franziska,Cairns, Timothy,Boecker, Simon,et al. Tet-on, or Tet-off, that is the question: Advanced conditional gene expression in Aspergillus[J]. FUNGAL GENETICS AND BIOLOGY,2016,89:72-83.
APA	Wanka, Franziska.,Cairns, Timothy.,Boecker, Simon.,Berens, Christian.,Happel, Anna.,...&Meyer, Vera.(2016).Tet-on, or Tet-off, that is the question: Advanced conditional gene expression in Aspergillus.FUNGAL GENETICS AND BIOLOGY,89,72-83.
MLA	Wanka, Franziska,et al."Tet-on, or Tet-off, that is the question: Advanced conditional gene expression in Aspergillus".FUNGAL GENETICS AND BIOLOGY 89(2016):72-83.