title: Enhanced Cry1Da production in Bacillus thuringiensis by driving expression from the σ(E) -dependent BtI promoter.
abstract: OBJECTIVE
  To increase the Cry1Da production in Bacillus thuringiensis by enhancing BtI promoter activity and fusion with upstream sequence from cry1Ab.
  
  
  RESULTS
  The effects of joining the upstream sequence of cry1Ab that contains E2 subunit pyruvate dehydrogenase (PDH) recognition site to the cry1Da promoter as well as the effects of substitution mutation of conserved sequences of its BtI promoter on cry1Da expression was monitored by constructing cry1Da promoter-lacZ fusions. Changing the -35 region of the cry1Da BtI promoter to that of cry1Ab enhanced β-galactosidase activity about three fold as comparing to that of the wild-type promoter with its own upstream sequence. In contrast, the same cry1Da mutated promoter linked to the above upstream sequence of cry1Ab enhanced enzyme activity up to seven fold, but was five fold lower than that of the full-length cry1Ab promoter. The cry1Ab-cry1Da hybrid promoter with the -35 BtI mutation efficiently increased Cry1Da synthesis by 133% and resulted in a 2·3-fold increase in insect larval toxicity when comparing to the wild type.
  
  
  CONCLUSIONS
  The cry1Ab promoter as well as mutation of -35 region of BtI promoter together with fusion with E2 subunit PDH recognition site efficiently enhanced Cry1Da production in B. thuringiensis.
  
  
  CONCLUSIONS
  The results provide useful information to construct an efficient cry1Da gene expression in B. thuringiensis.
authors.affiliation: Department of Biotechnology, Faculty of Science, Mahidol University, Bangkok, Thailand.
authors.forename: P
authors.initials: P
authors.lastname: Wanapaisan
authors.forename: O
authors.initials: O
authors.lastname: Chumsakul
authors.forename: W
authors.initials: W
authors.lastname: Panbangred
mesh_terms: D001413:Bacillus thuringiensis / Q000235:genetics* / Q000378:metabolism; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics*; D001483:Base Sequence; D051047:Dihydrolipoyllysine-Residue Acetyltransferase / Q000235:genetics; D004731:Endotoxins / Q000096:biosynthesis / Q000235:genetics*; D006460:Hemolysin Proteins / Q000096:biosynthesis / Q000235:genetics*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012808:Sigma Factor / Q000378:metabolism; D001616:beta-Galactosidase / Q000032:analysis / Q000235:genetics
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D012808:Sigma Factor; C030956:insecticidal crystal protein, Bacillus Thuringiensis; C063202:sporulation-specific sigma factors; D051047:Dihydrolipoyllysine-Residue Acetyltransferase; D001616:beta-Galactosidase
keywords: Bacillus thuringiensis; cry1Da; gene expression; σE-dependent promoter; σK-dependent promoter