Application of the bacteriophage Mu-driven system for the integration/amplification of target genes in the chromosomes of engineered Gram-negative bacteria-mini review

作者：Akhverdyan Valerii Z; Gak Evgueni R; Tokmakova Irina L; Stoynova Nataliya V; Yomantas Yurgis A V; Mashko Sergey V^*

来源：Applied Microbiology and Biotechnology, 2011, 91(4): 857-871.

DOI：10.1007/s00253-011-3416-y

• Chromosomal editing
• Dual-component system
• Enhancer element
• Excisable marker
• L/R ends
• MuA transposase
• Plasmid-less recombinant strain
• Replicative transposition

The advantages of phage Mu transposition-based systems for the chromosomal editing of plasmid-less strains are reviewed. The cis and trans requirements for Mu phage-mediated transposition, which include the L/R ends of the Mu DNA, the transposition factors MuA and MuB, and the cis/trans functioning of the E element as an enhancer, are presented. Mini-Mu(LR)/(LER) units are Mu derivatives that lack most of the Mu genes but contain the L/R ends or a properly arranged E element in cis to the L/R ends. The dual-component system, which consists of an integrative plasmid with a mini-Mu and an easily eliminated helper plasmid encoding inducible transposition factors, is described in detail as a tool for the integration/amplification of recombinant DNAs. This chromosomal editing method is based on replicative transposition through the formation of a cointegrate that can be resolved in a recombination-dependent manner. (E-plus)- or (E-minus)-helpers that differ in the presence of the trans-acting E element are used to achieve the proper mini-Mu transposition intensity. The systems that have been developed for the construction of stably maintained mini-Mu multi-integrant strains of Escherichia coli and Methylophilus methylotrophus are described. A novel integration/amplification/fixation strategy is proposed for consecutive independent replicative transpositions of different mini-Mu(LER) units with "excisable" E elements in methylotrophic cells.

• 出版日期2011-8