title: Construction of the plasmid-free strain for human growth hormone production.
abstract: The E. coli strain, overproducing human growth hormone (hGH) was made by integration of the hGH gene under the control of T7 promoter into the chromosomal LacZ gene of BL21(DE3) via lambda Red recombineering. The strain gave higher productivity (50 mg·L(-1)·OD550(-1)) and better growth characteristics than the corresponding strain in which the same hGH expression cassette was placed in a plasmid. The protein produced by the plasmid-free strain was purified and characterized to be hGH. The results demonstrates that a plasmid-free recombinant strain having a single-copy gene expression cassette in the chromosome could provide better gene activity regulation, higher productivity, superior growth characteristics, as well as more stringent control of the gene sequence invariance than a plasmid-based strain.
authors.affiliation: Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia.
authors.forename: A A
authors.initials: AA
authors.lastname: Schulga
authors.affiliation: Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia.
authors.forename: P V
authors.initials: PV
authors.lastname: Mechev
authors.affiliation: Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia; Lomonosov Moscow State University, Faculty of Biology, 119991 Moscow, Russia.
authors.forename: M P
authors.initials: MP
authors.lastname: Kirpichnikov
authors.affiliation: Institute of Bioengineering, Russian Academy of Sciences, 117312 Moscow, Russia.
authors.forename: K G
authors.initials: KG
authors.lastname: Skryabin
authors.affiliation: Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia. Electronic address: deyev@ibch.ru.
authors.forename: S M
authors.initials: SM
authors.lastname: Deyev
mesh_terms: D017123:Bacteriophage T7 / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics*; D005786:Gene Expression Regulation; D019382:Human Growth Hormone / Q000096:biosynthesis* / Q000235:genetics*; D006801:Humans; D007763:Lac Operon / Q000235:genetics; D013058:Mass Spectrometry / Q000379:methods; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification
chemical_list: D011994:Recombinant Proteins; D019382:Human Growth Hormone
keywords: Human growth hormone; Lambda Red recombineering; Plasmid-free; Somatotropin