Automatic elimination of unnecessary bacterial sequences from yeast vectors

• Published in: Gene Vol. 121; no. 1; pp. 161 - 165
• Main Authors: Kaoru, Awane, Akira, Naito, Hiroyuki, Araki, Oshima, Yasuji
• Format: Journal Article
• Language: English
• Published: Lausanne Elsevier B.V 02.11.1992; Amsterdam Elsevier; New York, NY
• Subjects: Biological and medical sciences; Biotechnology; DNA, Bacterial; Escherichia coli - genetics; Fundamental and applied biological sciences. Psychology; Gene Conversion; Genetic engineering; Genetic technics; Genetic Vectors; Methods. Procedures. Technologies; plasmid stability; Plasmids; pSR1 plasmid; Recombination, Genetic; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Sequence Deletion; site-specific recombination; Vectors (cloning, transfer, expression). Insertion sequences and transposons; plasmid stability; ORF; E; nt; pSR1 plasmid; IR; bp; Ap; s; kb; site-specific recombination; Z; SD-Leu(medium); Gene conversion; Saccharomyces cerevisiae; Yeast; Escherichia coli; Nucleotide fragment; Fungi; Deletion; Shuttle vector; Bacteria; Ascomycetes; Performance analysis; Enterobacteriaceae; Thallophyta
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/0378-1119(92)90176-P

Most vectors for Saccharomyces cerevisiae are shuttle vectors which can be both propagated and selected in Escherichia coli. The DNA segments, however, which are required for propagation in E. coli are unnecessary and moreover toxic in S. cerevisiae. To delete these harmful DNA fragments from the vector after it is introduced into S. cerevisiae cells, we propose a specific gene conversion mechanism of a yeast plasmid, pSRl. Plasmid pSRl has a pair of inverted repeats (IRs) that divides the plasmid molecule into two unique regions. Intramolecular recombination frequently occurs at a pair of specific recombination sites in IRs catalyzed by recombinase R, encoded by a pSRl plasmid gene. This R-mediated recombination is often accompanied by gene conversion in IRs. Thus, a 2.1-kb pBR322 sequence for the E. coli host ligated into one of the IRs of a composite plasmid was automatically and effectively eliminated when the plasmid was introduced into S. cerevisiae cells.