TABLE 1.

Bacterial strains and plasmids

Bacterial strain or plasmidRelevant characteristic(s)aReference or source
Strain
    E. coli ER2275NEBb
    E. coli OneShot chemically     competent TOP10Invitrogen
    C. acetobutylicum ATCC 824ATCCc
    M5AADC CoAT4
Plasmid
    pAN1Cmr; carries the φ3T I gene17
    pFNK6Ampr MLSr; ace operon18
    pFNK7Ampr MLSr ctfA ctfB18
    pSOS95ace2 operon30
    pSOS95delAmpr MLSr; thl promoterThis study
    pADC38ASAmpr MLSr; thl promoter; antisense adc38This study
    pADC68ASAmpr MLSr; thl promoter; antisense adc68This study
    pADC100ASAmpr MLSr; thl promoter; antisense adc100This study
    pCTFA2ASAmpr MLSr; thl promoter; antisense ctfa2This study
    pCTFB1ASAmpr MLSr; thl promoter, antisense ctfb1This study
    pCOAT11ASAmpr MLSr; thl promoter; combined antisense ctfb1 and antisense ctfa1This study
  • a adc, AADC gene; ctfA, CoAT subunit A gene; ctfB, CoAT subunit B gene; Cmr, chloramphenicol resistant; φ3T I, Bacillus subtilis phage φ3T I methyltransferase gene; Ampr, ampicillin resistant; MLSr, macrolide-lincosamide-streptogramin B resistant; ace operon, synthetic operon which contains the three homologous acetone formation genes (adc, ctfA, and ctfB) transcribed from the adc promoter; ctfA, gene encoding for the A subunit of CoAT; ctfB, gene encoding for the B subunit of CoAT; ace2 operon, synthetic operon which contains the three homologous acetone formation genes (adc, ctfA, and ctfB) transcribed from the thl promoter; thl promoter, promoter region for the thiolase gene (thl) of C. acetobutylicum strain ATCC 824; adc38, DNA fragment containing approximately 38% of the sequence of the adc transcript; adc68, DNA fragment containing approximately 68% of the sequence of the adc transcript; adc100, DNA fragment containing approximately 100% of the sequence of the adc transcript; ctfa2, DNA fragment containing approximately 79% of the ctfA structural gene; ctfb1, DNA fragment containing approximately 39% of the ctfB structural gene; ctfa1, DNA fragment containing approximately 53% of the ctfA structural gene.

  • b NEB, New England Biolabs (Beverly, Mass.).

  • c ATCC, American Type Culture Collection (Manassas, Va.).