Ann N Y Acad Sci. 1992 Oct 13;665:39-51.

Vector construction, transformation, and gene amplification in Clostridium acetobutylicum ATCC 824.

Lee SY, Mermelstein LD, Bennett GN, Papoutsakis ET.

Source

Department of Chemical Engineering, Northwestern University, Evanston, Illinois 60208.

Abstract

In order to alter the primary metabolism of C. acetobutylicum, we have constructed E. coli- or B. subtilis-C. acetobutylicum shuttle vectors that could be used to deliver homologous fermentative genes into C. acetobutylicum ATCC 824. The plasmid copy number and plasmid stability in C. acetobutylicum for several of these plasmids were determined. We have also developed a protocol for the electrotransformation of C. acetobutylicum ATCC 824. Difficulty in the transformation of C. acetobutylicum ATCC 824 with vectors containing DNA from E. coli plasmids was found to be due to the existence of a restriction system in this strain. This type II restriction endonuclease, named Cac824I, recognizes the sequence 5'-GCNGC-3' and cuts ColE1 plasmids frequently. One of the vectors, pFNK1, possessing a variety of unique cloning sites was used in the amplification of one acid (PTB) and one solvent (AADC) formation gene. The corresponding enzyme activities were amplified in C. acetobutylicum as shown by enzyme assays and SDS-PAGE gels of cell extracts.

PMID:: 1416617; [PubMed - indexed for MEDLINE]

Publication Types, MeSH Terms, Substances

Publication Types

Research Support, U.S. Gov't, Non-P.H.S.

MeSH Terms

Substances

DNA, Bacterial

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Cited by 5 PubMed Central articles

Inactivation of σF in Clostridium acetobutylicum ATCC 824 blocks sporulation prior to asymmetric division and abolishes σE and σG protein expression but does not block solvent formation. [J Bacteriol. 2011]
Inactivation of σF in Clostridium acetobutylicum ATCC 824 blocks sporulation prior to asymmetric division and abolishes σE and σG protein expression but does not block solvent formation.
Jones SW, Tracy BP, Gaida SM, Papoutsakis ET. J Bacteriol. 2011 May; 193(10):2429-40. Epub 2011 Mar 18.
Inactivation of σE and σG in Clostridium acetobutylicum illuminates their roles in clostridial-cell-form biogenesis, granulose synthesis, solventogenesis, and spore morphogenesis. [J Bacteriol. 2011]
Inactivation of σE and σG in Clostridium acetobutylicum illuminates their roles in clostridial-cell-form biogenesis, granulose synthesis, solventogenesis, and spore morphogenesis.
Tracy BP, Jones SW, Papoutsakis ET. J Bacteriol. 2011 Mar; 193(6):1414-26. Epub 2011 Jan 7.
Engineering clostridium strain to accept unmethylated DNA. [PLoS One. 2010]
Engineering clostridium strain to accept unmethylated DNA.
Dong H, Zhang Y, Dai Z, Li Y. PLoS One. 2010 Feb 9; 5(2):e9038. Epub 2010 Feb 9.

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