title: Gene silencing in Escherichia coli using antisense RNAs expressed from doxycycline-inducible vectors.
abstract: Here, we report on the construction of doxycycline (tetracycline analogue)-inducible vectors that express antisense RNAs in Escherichia coli. Using these vectors, the expression of genes of interest can be silenced conditionally. The expression of antisense RNAs from the vectors was more tightly regulated than the previously constructed isopropyl-β-D-galactopyranoside-inducible vectors. Furthermore, expression levels of antisense RNAs were enhanced by combining the doxycycline-inducible promoter with the T7 promoter-T7 RNA polymerase system; the T7 RNA polymerase gene, under control of the doxycycline-inducible promoter, was integrated into the lacZ locus of the genome without leaving any antibiotic marker. These vectors are useful for investigating gene functions or altering cell phenotypes for biotechnological and industrial applications.
  
  
  CONCLUSIONS
  A gene silencing method using antisense RNAs in Escherichia coli is described, which facilitates the investigation of bacterial gene function. In particular, the method is suitable for comprehensive analyses or phenotypic analyses of genes essential for growth. Here, we describe expansion of vector variations for expressing antisense RNAs, allowing choice of a vector appropriate for the target genes or experimental purpose.
authors.affiliation: Bioproduction Research Institute, National Institute of Advanced Industrial Sciences and Technology AIST, Sapporo 062-8517, Japan. n-nakashima@aist.go.jp
authors.forename: N
authors.initials: N
authors.lastname: Nakashima
authors.forename: T
authors.initials: T
authors.lastname: Tamura
mesh_terms: D000103:Acetyl-CoA Carboxylase / Q000235:genetics; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D004318:Doxycycline / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development; D029968:Escherichia coli Proteins / Q000235:genetics; D054889:Fatty Acid Synthase, Type II / Q000235:genetics; D020868:Gene Silencing; D005798:Genes, Bacterial; D005822:Genetic Vectors; D007544:Isopropyl Thiogalactoside / Q000031:analogs & derivatives; D016372:RNA, Antisense / Q000235:genetics / Q000378:metabolism; D012329:RNA, Bacterial / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics
chemical_list: D029968:Escherichia coli Proteins; D016372:RNA, Antisense; D012329:RNA, Bacterial; D014764:Viral Proteins; D007544:Isopropyl Thiogalactoside; C070641:isopropyl-beta-galactopyranoside; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D054889:Fatty Acid Synthase, Type II; D000103:Acetyl-CoA Carboxylase; C557124:accA protein, E coli; D004318:Doxycycline