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Plant and Cell Physiology, 1986, Vol. 27, No. 5 801-808
© 1986

Article

Stepwise Transformation in Saccharomyces cerevisiae Yeast: Construction of Strains for Transformation and Subsequent Cytoductive Transfer of Plasmid DNA with Mitochondria1

Katsunori Suzuki and Kazuo Yoshida2

Biological Institute, Faculty of Science, Nagoya University Nagoya, 464, Japan

2 To whom reprint requests should be addressed.

As a general transformation method in Saccharomyces cerevisiae, a "stepwise transformation method" of three steps was developed: conventional transformation of established carrier strains with plasmid DNA, subsequent effective transfer of plasmid to destination hosts either by cytoduction or by meiosis after mating, and finally a simple test for plasmid retention. The several auxotrophic carrier strains constructed were readily transformable, defective in karyogamy (kar 1), and highly susceptible to cell-wall lytic enzymes. Effective transfer of plasmid DNA from the carrier strains to other strains by cytoduction or meiosis was confirmed, and the target transformants were obtained. The simple method for checking plasmid retention was based on the cytoductive transfer of plasmid DNA to tester cells which have recessive multiple drug resistance and kar 1 mutations. The stepwise method should largely extend the range of strains which can be transformed and spare time and labor. Its limitations as well as other advantages are also discussed.

1 The preliminary report of this paper was presented in the Annual Meeting of the Japanese Society of Plant Physiologists, Kanazawa, April, 1984.

(Received November 5, 1985; Accepted April 7, 1986)
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