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Journal of Bacteriology, March 2002, p. 1449-1454, Vol. 184, No. 5
0021-9193/02/$04.00+0     DOI: 10.1128/JB.184.5.1449-1454.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Directed Mutagenesis and Plasmid-Based Complementation in the Methanogenic Archaeon Methanosarcina acetivorans C2A Demonstrated by Genetic Analysis of Proline Biosynthesis

*** Jun Kai Zhang, Andrea K. White, H. Craig Kuettner, Paolo Boccazzi, and William W. Metcalf*

Department of Microbiology, University of Illinois, Urbana, Illinois 61801

Received 7 September 2001/ Accepted 28 November 2001

We report here the first use of directed mutagenesis in Methanosarcina acetivorans C2A. The method employs homologous recombination-mediated gene replacement and was used to construct a variety of proline auxotrophs with mutations in the proABC locus. Each mutation was also complemented in trans with autonomously replicating Methanosarcina-Escherichia plasmid shuttle vectors.

* Corresponding author. Mailing address: Department of Microbiology, University of Illinois, B103 Chemical and Life Science Laboratory, 601 S. Goodwin Ave., Urbana, IL 61801. Phone: (217) 244-1943. Fax: (217) 244-6697. E-mail: metcalf{at}uiuc.edu.

Journal of Bacteriology, March 2002, p. 1449-1454, Vol. 184, No. 5
0021-9193/02/$04.00+0     DOI: 10.1128/JB.184.5.1449-1454.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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