title: Application of the bacteriophage Mu-driven system for the integration/amplification of target genes in the chromosomes of engineered Gram-negative bacteria--mini review.
abstract: 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.
authors.affiliation: Ajinomoto-Genetika Research Institute, Moscow 117545, Russian Federation.
authors.forename: Valerii Z
authors.initials: VZ
authors.lastname: Akhverdyan
authors.forename: Evgueni R
authors.initials: ER
authors.lastname: Gak
authors.forename: Irina L
authors.initials: IL
authors.lastname: Tokmakova
authors.forename: Nataliya V
authors.initials: NV
authors.lastname: Stoynova
authors.forename: Yurgis A V
authors.initials: YA
authors.lastname: Yomantas
authors.forename: Sergey V
authors.initials: SV
authors.lastname: Mashko
mesh_terms: D010583:Bacteriophage mu / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D005827:Genetics, Microbial / Q000379:methods*; D020630:Methylophilus methylotrophus / Q000235:genetics*; D016254:Mutagenesis, Insertional / Q000379:methods*; D011995:Recombination, Genetic