TY - JOUR
T1 - Evaluation of chromosomal insertion loci in the Pseudomonas putida KT2440 genome for predictable biosystems design
AU - Chaves, Julie E.
AU - Wilton, Rosemarie
AU - Gao, Yuqian
AU - Munoz, Nathalie Munoz
AU - Burnet, Meagan C.
AU - Schmitz, Zachary
AU - Rowan, John
AU - Burdick, Leah H.
AU - Elmore, Joshua
AU - Guss, Adam
AU - Close, Dan
AU - Magnuson, Jon K.
AU - Burnum-Johnson, Kristin E.
AU - Michener, Joshua K.
N1 - Publisher Copyright:
© 2020 The Authors
PY - 2020/12
Y1 - 2020/12
N2 - The development of Pseudomonas strains for industrial production of fuels and chemicals will require the integration of heterologous genes and pathways into the chromosome. Finding the most appropriate integration site to maximize strain performance is an essential part of the strain design process. We characterized seven chromosomal loci in Pseudomonas putida KT2440 for integration of a fluorescent protein expression construct. Insertion in five of the loci did not affect growth rate, but fluorescence varied by up to 27-fold. Three sites displaying a diversity of phenotypes with the fluorescent reporter were also chosen for the integration of a gene encoding a muconate importer. Depending on the integration locus, expression of the importer varied by approximately 3-fold and produced significant phenotypic differences. This work demonstrates the impact of the integration location on host viability, gene expression, and overall strain performance.
AB - The development of Pseudomonas strains for industrial production of fuels and chemicals will require the integration of heterologous genes and pathways into the chromosome. Finding the most appropriate integration site to maximize strain performance is an essential part of the strain design process. We characterized seven chromosomal loci in Pseudomonas putida KT2440 for integration of a fluorescent protein expression construct. Insertion in five of the loci did not affect growth rate, but fluorescence varied by up to 27-fold. Three sites displaying a diversity of phenotypes with the fluorescent reporter were also chosen for the integration of a gene encoding a muconate importer. Depending on the integration locus, expression of the importer varied by approximately 3-fold and produced significant phenotypic differences. This work demonstrates the impact of the integration location on host viability, gene expression, and overall strain performance.
UR - http://www.scopus.com/inward/record.url?scp=85088999458&partnerID=8YFLogxK
U2 - 10.1016/j.mec.2020.e00139
DO - 10.1016/j.mec.2020.e00139
M3 - Article
AN - SCOPUS:85088999458
SN - 2214-0301
VL - 11
JO - Metabolic Engineering Communications
JF - Metabolic Engineering Communications
M1 - e00139
ER -