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Centromeric DNA of Kluyveromyces lactis
J J Heus1, B J Zonneveld, H Y Steensma
1Department of Cell Biology and Genetics, Leiden University, The Netherlands.
Current Genetics
|December 1, 1990
Summary
Researchers isolated five centromeres from the yeast Kluyveromyces lactis using a novel direct selection method. These isolated centromeres confer mitotic stability but do not function in Saccharomyces cerevisiae.
Area of Science:
- Molecular Biology
- Yeast Genetics
- Chromosomal Biology
Background:
- Centromeres are crucial for chromosome segregation during cell division.
- Understanding centromere function requires methods to isolate and study these regions.
- Kluyveromyces lactis presents a model system for studying eukaryotic centromeres.
Purpose of the Study:
- To develop and apply a direct selection method for isolating centromeres from Kluyveromyces lactis.
- To characterize the isolated centromeric fragments and their functional properties.
Main Methods:
- Direct selection method utilizing the lethality of the SUP11 tRNA gene at high copy number.
- Construction of plasmids with a Kluyveromyces lactis replication origin (KARS) and isolated fragments.
- Assessment of plasmid mitotic stability and copy number.
- Chromosomal hybridization to confirm fragment origin.
Main Results:
- Successfully isolated five distinct chromosomal fragments conferring mitotic stability to KARS plasmids.
- KARS plasmids with these fragments exhibited a low copy number (approximately one).
- Each fragment hybridized to a different chromosome, indicating unique centromeric origins.
- Isolated centromeres did not function in the budding yeast Saccharomyces cerevisiae.
Conclusions:
- Five of the six centromeres of Kluyveromyces lactis were successfully isolated using the described method.
- The isolated centromeres are functional in K. lactis, ensuring mitotic stability.
- The isolated centromeres are species-specific and do not function in Saccharomyces cerevisiae, highlighting differences in centromere recognition or structure.
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