title: [Construction of Escherichia coli-Streptomyces shuttle expression plasmid pMF].
abstract: OBJECTIVE
  To construct an E. coli-streptomyces shuttle vector pMF that can integrate into the genome of streptomyces by site-specific integration.
  
  
  METHODS
  We inserted the integrase gene phiC31 int and attP site into pLSB2, a suicidal streptomyces plasmid. The resulting conjugably transferable vector which contains the activator promoter system act II-ORF4/Pact I from Strepromyces coelicolor A3 (2) could be integrated into the genome of streptomyces by site-specific integration.
  
  
  RESULTS
  The plasmid pMF was conjugably transferred with high frequency into S. coelicolor M145, S. lividans TK24 and Saccharopolyspora erythraea 2338 from E. coll. Southern blotting results showed that pMF was able to integrate into the genome of streptomyces. We also confirmed functional protein expression by cloning a putative S-adenosylmethionine synthetase (SAM-s) gene from Sacc. spinosa S08-4 into pMF and conjugated into S. coelicolor M145. Protein expression were confirmed using Western blotting.
  
  
  CONCLUSIONS
  pMF can be used as an effective tool for site-specific integration expression of foreign gene in streptomyces.
authors.affiliation: Key Lab of Microbial Molecular Biology of Hunan Province, College of Life Science, Hunan Normal University, Changsha 410081, China. quanmeifang@yeah.net
authors.forename: Meifang
authors.initials: M
authors.lastname: Quan
authors.forename: Kai
authors.initials: K
authors.lastname: Yu
authors.forename: Liqiu
authors.initials: L
authors.lastname: Xia
authors.forename: Xuezhi
authors.initials: X
authors.lastname: Ding
authors.forename: Fanjun
authors.initials: F
authors.lastname: Zeng
authors.forename: Xiaoxiao
authors.initials: X
authors.lastname: Kou
authors.forename: Hailong
authors.initials: H
authors.lastname: Wang
authors.forename: Shengbiao
authors.initials: S
authors.lastname: Hu
authors.forename: Ziquan
authors.initials: Z
authors.lastname: Yu
authors.forename: Jingye
authors.initials: J
authors.lastname: Li
mesh_terms: D003227:Conjugation, Genetic / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D010957:Plasmids / Q000235:genetics*; D013302:Streptomyces / Q000235:genetics*