PMID: 15974564Jun 25, 2005Paper

Visualization of protoplast fusion and quantitation of recombination in fused protoplasts of auxotrophic strains of Escherichia coli

Metabolic Engineering
MingHua DaiShelley D Copley

Abstract

Protoplast fusion has been used to combine genes from different organisms to create strains with desired properties. A recently developed variant on this approach, genome shuffling, involves generation of a genetically heterogeneous population of a single organism, followed by recursive protoplast fusion to allow recombination of mutations within the fused protoplasts. These are powerful techniques for engineering of microbial strains for desirable industrial properties. However, there is a prevailing opinion that it will be difficult to use these methods for engineering of Gram-negative bacteria because the outer membrane makes protoplast fusion more difficult. Here we describe the successful use of protoplast fusion in Escherichia coli. Using two auxotrophic strains of E. coli, we obtained prototrophic strains by recombination in fused protoplasts at frequencies of 0.05-0.7% based on the number of protoplasts subjected to fusion. This frequency is three-four orders of magnitude better than those previously reported for recombination in fused protoplasts of Gram-negative bacteria such as E. coli and Providencia alcalifaciens.

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Citations

Sep 18, 2013·Applied Biochemistry and Biotechnology·Wei LiZhiqun Liang
Mar 7, 2014·Applied Microbiology and Biotechnology·Damien Biot-Pelletier, Vincent J J Martin
May 17, 2011·Microbial Cell Factories·Tomohiro MakinoGeorge Georgiou
Nov 2, 2012·Antimicrobial Agents and Chemotherapy·Christopher P RandallAlex J O'Neill
Mar 30, 2010·Biotechnology Letters·James WinklerKaty C Kao
Jul 24, 2010·Journal of Industrial Microbiology & Biotechnology·Dao-Qiong ZhengYu-Hua Zhao
Jul 31, 2012·Metabolic Engineering·James Winkler, Katy C Kao
May 26, 2009·Biotechnology Advances·Jixian GongXueming Zhao
Sep 1, 2015·Current Opinion in Biotechnology·Kyle A Zingaro, Eleftherios Terry Papoutsakis
Jan 1, 2012·Computational and Structural Biotechnology Journal·Georgios Skretas, Fragiskos N Kolisis
Jan 31, 2015·Applied Microbiology and Biotechnology·Wenshan Liu, Rongrong Jiang
Mar 6, 2019·Life·Kazuhito V TabataHiroyuki Noji
Jan 19, 2020·Experimental Cell Research·David M Brown, John I Glass

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