Expression and identification of immunity determinants on linear DNA killer plasmids pGKL1 and pGKL2 in Kluyveromyces lactis.
نویسندگان
چکیده
The linear dsDNA plasmids, pGKL1 (8.9 kb) and pGKL2 (13.4 kb) discovered in Kluyveromyces lactis, confer killer and immunity characteristics upon various yeast strains. We have devised an immunity assay and have been able to show the expression of an immunity phenotype in the K. lactis transformants harbouring conventional circular plasmids which contain DNA fragments of pGKL1. Using this expression system, the immunity determinant on pGKL1 was identified as ORF5. In addition, the presence of pGKL2 was proved to be essential for the expression of the immunity phenotype. This is the first demonstration of this new pGKL2 function, as distinct from its known functions for the replication and maintenance of pGKL1 in yeast cells.
منابع مشابه
Efficient isolation of the linear DNA killer plasmid of Kluyveromyces lactis: evidence for location and expression in the cytoplasm and characterization of their terminally bound proteins.
Differential centrifugation of an osmotic lysate of K. lactis protoplasts showed that the linear DNA killer plasmids of K. lactis, pGKL1 and pGKL2, are almost exclusively present in the cytoplasmic fraction. This fractionation procedure allows the rapid isolation of large amounts of plasmid DNA without contamination by chromosomal and mitochondrial DNA. With these DNA preparations the size of t...
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Linear DNA plasmids were found in the following yeasts: four strains of Kluyveromyces lactis, one of Debaryomyces hansenii, one of Wingea robertsiae and four of Pichia etchellsii. In each case, the plasmids were present as a pair of DNA molecules of different sizes. The plasmids of K. lactis strains were associated with a killer activity and their structure was similar to the known killer plasm...
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متن کاملThe yeast linear DNA killer plasmids, pGKL1 and pGKL2, possess terminally attached proteins.
The terminal structures of linear DNA killer plasmids from yeast, pGKL1 and pGKL2, were analyzed. Results obtained by exonuclease treatments of these plasmids show that both pGKL plasmids have free hydroxyl 3'-ends and blocked 5'-ends. Electrophoretic analysis of the terminal restriction fragments treated with proteases revealed that pGKL1 and pGKL2 have proteins bound at 5'termini and that the...
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ورودعنوان ژورنال:
- Nucleic acids research
دوره 15 3 شماره
صفحات -
تاریخ انتشار 1987