title: Flagellar region 3b supports strong expression of integrated DNA and the highest chromosomal integration efficiency of the Escherichia coli flagellar regions.
abstract: The Gram-negative bacterium Escherichia coli is routinely used as the chassis for a variety of biotechnology and synthetic biology applications. Identification and analysis of reliable chromosomal integration and expression target loci is crucial for E. coli engineering. Chromosomal loci differ significantly in their ability to support integration and expression of the integrated genetic circuits. In this study, we investigate E. coli K12 MG1655 flagellar regions 2 and 3b. Integration of the genetic circuit into seven and nine highly conserved genes of the flagellar regions 2 (motA, motB, flhD, flhE, cheW, cheY and cheZ) and 3b (fliE, F, G, J, K, L, M, P, R), respectively, showed significant variation in their ability to support chromosomal integration and expression of the integrated genetic circuit. While not reducing the growth of the engineered strains, the integrations into all 16 target sites led to the loss of motility. In addition to high expression, the flagellar region 3b supports the highest efficiency of integration of all E. coli K12 MG1655 flagellar regions and is therefore potentially the most suitable for the integration of synthetic genetic circuits.
authors.affiliation: Department of Pathology, University of Cambridge, Tennis Court Road, CB2 1QP, Cambridge, UK.
authors.forename: Mario
authors.initials: M
authors.lastname: Juhas
authors.affiliation: Department of Pathology, University of Cambridge, Tennis Court Road, CB2 1QP, Cambridge, UK.
authors.forename: James W
authors.initials: JW
authors.lastname: Ajioka
mesh_terms: D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development / Q000502:physiology; D005407:Flagella / Q000235:genetics* / Q000502:physiology; D015870:Gene Expression; D056426:Genetic Loci; D008124:Locomotion; D060847:Metabolic Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D011995:Recombination, Genetic
chemical_list: D011994:Recombinant Proteins