Operation of an efficient site-specific recombination system of Zygosaccharomyces rouxii in tobacco cells (original) (raw)

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Department of Fermentation Technology, Faculty of Engineering, Osaka University

2-1 Yamadaoka, suita-shi, Osaka 565, Japan

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Department of Fermentation Technology, Faculty of Engineering, Osaka University

2-1 Yamadaoka, suita-shi, Osaka 565, Japan

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Faculty of Agriculture, Nagoya University

Chikusa-ku, Nagoya 464-01

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Faculty of Agriculture, Nagoya University

Chikusa-ku, Nagoya 464-01

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Received:

01 October 1991

Accepted:

13 November 1991

Published:

11 December 1991

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Hitoshi Onouchi, Kumi Yokoi, Chiyoko Machida, Hiroaki Matsuzaki, Yasuji Oshima, Ken Matsuoka, Kenzo Nakamura, Yasunori Machida, Operation of an efficient site-specific recombination system of Zygosaccharomyces rouxii in tobacco cells , Nucleic Acids Research, Volume 19, Issue 23, 11 December 1991, Pages 6373–6378, https://doi.org/10.1093/nar/19.23.6373
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Abstract

Recombinase encoded by the R gene of pSR1 of Zygosaccharomyces rouxii mediates reciprocal recombination between two specific recombination sites (RSs) to induce excision or inversion of the DNA segment that is flanked by the RSs. We report here that site-specific recombination mediated by this system takes place efficiently in tobacco cells. To monitor the recombination events in tobacco cells, we have constructed two types of cryptic β-glucuronidase reporter gene in such a way that recombination such as inversion of the construct or excision of the intervening sequence results in their expression. When these cryptic reporter constructs were transiently introduced together with the R gene by electroporation into protoplasts of tobacco cells, β-glucuronidase activity was detected. The cryptic reporter genes, when stably resident in the chromosome of tobacco cells, were also activated by the R gene. Structural analyses of the genomic DNA isolated from these tobacco cells showed that the R protein did in fact catalyze precise recombination between two copies of RSs in tobacco cells, with resultant activation of the cryptic reporter genes. This observation provides the basis for development of a DNA technology whereby large regions of DNA can be manipulated in plant chromosomes. Potential uses of this recombination system are discussed.

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© 1991 Oxford University Press

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