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2-micrometers DNA-like plasmids in the osmophilic haploid yeast Saccharomyces rouxii
Abstract:
DNA plasmids were detected in two independent strains of Saccharomyces rouxii among 100 yeast strains other than Saccharomyces cerevisiae tested. The plasmids, pSR1 and pSR2, had almost the same mass (approximately 4 X 10(6) daltons) as 2-micrometers DNA of S. cerevisiae. pSR1 and pSR2 gave identical restriction maps with restriction endonucleases BamHI, EcoRI, HincII, HindIII, and XhoI, and both lacked restriction sites for PstI, SalI, and SmaI. These maps, however, differed significantly from that of S. cerevisiae 2-micrometers DNA. Restriction analysis also revealed two isomeric forms of each plasmid and suggested the presence of a pair of inverted repeat sequences in the molecules where intramolecular recombination took place. DNA-DNA hybridization between the pSR1 and pSR2 DNAs indicated significant homology between their base sequences, whereas no homology was detected between pSR1 and pJDB219, a chimeric plasmid constructed from a whole molecule of 2-micrometers DNA, plasmid pMB9, and a 1.2-kilobase DNA fragment of S. cerevisiae bearing the LEU2 gene. A chimeric plasmid constructed with pSR1 and YIp1, the larger EcoRI-SalI fragment of pBR322 ligated with a 6.1-kilobase DNA fragment of S. cerevisiae bearing the HIS3 gene, could replicate autonomously in an S. cerevisiae host and produced isomers, presumably by intramolecular recombination at the inverted repeats.
Insights
Novel DNA plasmids, pSR1 and pSR2, were discovered in Saccharomyces rouxii strains. These plasmids exhibit unique restriction maps and homologous sequences, distinct from Saccharomyces cerevisiae 2-micrometers DNA.
Area of Science:
- Yeast genetics
- Molecular biology
- Plasmid research
Background:
- Plasmids are extrachromosomal DNA elements found in various organisms, including yeast.
- The 2-micrometers DNA plasmid is a well-characterized element in Saccharomyces cerevisiae.
- Investigating plasmids in other yeast species can reveal novel genetic elements and mechanisms.
Purpose of the Study:
- To identify and characterize novel plasmids in yeast strains other than Saccharomyces cerevisiae.
- To compare the characteristics of newly discovered plasmids with known yeast plasmids like the 2-micrometers DNA.
- To investigate the potential for autonomous replication and recombination of these novel plasmids.
Main Methods:
- Screening of 100 yeast strains (excluding S. cerevisiae) for the presence of DNA plasmids.
- Restriction enzyme digestion and analysis to generate restriction maps.
- DNA-DNA hybridization to assess sequence homology.
- Construction of chimeric plasmids for functional analysis (replication and isomer formation).
Main Results:
- Two plasmids, pSR1 and pSR2, were detected in Saccharomyces rouxii strains.
- pSR1 and pSR2 share similar mass and identical restriction maps, distinct from S. cerevisiae 2-micrometers DNA.
- These plasmids possess inverted repeat sequences, enabling intramolecular recombination and forming isomers.
- No homology was found between pSR1 and the chimeric plasmid pJDB219.
- A chimeric plasmid containing pSR1 demonstrated autonomous replication in S. cerevisiae, producing isomers.
Conclusions:
- Saccharomyces rouxii harbors unique plasmids, pSR1 and pSR2, with distinct genetic features compared to S. cerevisiae 2-micrometers DNA.
- The presence of inverted repeats suggests a mechanism for intramolecular recombination and plasmid variation.
- These novel plasmids have the potential for autonomous replication in heterologous yeast hosts, indicating functional similarities to known yeast plasmids.