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Journal of Bacteriology, September 2001, p. 5453-5458, Vol. 183, No. 18
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.18.5453-5458.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Recombinational Transfer of 100-Kilobase Genomic DNA to Plasmid in Bacillus subtilis 168

Kenji Tsuge and Mitsuhiro Itaya*

Mitsubishi Kasei Institute of Life Sciences, Machida-Shi, Tokyo 194-8511, Japan

Received 13 April 2001/Accepted 26 June 2001

Transformation of Bacillus subtilis by a plasmid requires a circular multimeric form. In contrast, linearized plasmids can be circularized only when homologous sequences are present in the host genome. A recombinational transfer system was constructed with this intrinsic B. subtilis recombinational repair pathway. The vector, pGETS103, a derivative of the theta -type replicating plasmid pTB19 of thermophilic Bacillus, had the full length of Escherichia coli plasmid pBR322. A multimeric form of pGETS103 yielded tetracycline-resistant transformants of B. subtilis. In contrast, linearized pGETS103 gave tetracycline-resistant transformants only when the recipient strain had the pBR322 sequence in the genome. The efficiency and fidelity of the recombinational transfer of DNAs of up to 90 kb are demonstrated.

* Corresponding author. Mailing address: Mitsubishi Kasei Institute of Life Sciences, 11 Minamioya, Machida-Shi, Tokyo 194-8511, Japan. Phone: 81-42-724-6352. Fax: 81-42-724-6316. E-mail: ita{at}libra.ls.m-kagaku.co.jp.

Journal of Bacteriology, September 2001, p. 5453-5458, Vol. 183, No. 18
0021-9193/01/$04.00+0   DOI: 10.1128/JB.183.18.5453-5458.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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Copyright © 2001 by the American Society for Microbiology. All rights reserved.