Meiotic and Mitotic Phenotypes Conferred by the blm1-1 Mutation in Saccharomyces cerevisiae and MSH4 Suppression of the Bleomycin Hypersusceptibility
Abstract
:Introduction
Materials and Methods
Strains and Plasmids
Strain or Plasmid | Genotype or Selectable Markers | Source |
Saccharomyces cerevisiae | ||
CM1457-79A | MATα ade2-1 and/or ade2-40 trp1-1 and/or trp5b | This laboratory |
CM1457-79B | MATa ade2-1 and/or ade2-40 ilv1-92 trp1-1 and/or trp5b | This laboratory |
CM1457-79C | MATa ade2-1 and/or ade2-40 blm1-1 trp1-1 and/or trp5b leu2-3 and/or leu2-112 ura3-1 | This laboratory |
CM1457-79D | MATα ade2-1 and/or ade2-40 blm1-1 ilv1-92 leu2-3 and/or leu2-112 trp1-1 and/or trp5b ura3-1 | This laboratory |
PS593/6H | MATα his3Δ200 leu2-3 or leu2-112 msh4::LEU2 trp1-289 ura3-52 | Dr. Nancy Hollingsworth; Dr. Shirleen Roeder [15] |
Plasmids | ||
YCp50 | amp tet ARS1 CEN4 URA3 | Johnston and Davis, 1984 [16] |
pPM118 | tet ARS1 CEN4 URA3 | This laboratory |
pPM118-4 | tet ARS1 CEN4 URA3 | This laboratory |
Media and Growth Conditions
Sporulation and Dissection of Tetrads
Zeocin Treatments
E. coli and Yeast Transformations
Results
Reduced Viability of Spores from Homozygous Mutant Strains
Diploid Type | Total number of tetrads dissected | No viable spores | 1 viable spore | 2 viable spores | 3 viable spores | 4 viable spores |
Homozygous normal | 16 | 0 | 1 | 0 | 6 | 9 |
Heterozygous | 14 | 0 | 0 | 0 | 5 | 9 |
Homozygous mutant | 31 | 0 | 7 | 20 | 4 | 0 |
Diploid Type | Total number of tetrads dissected | Sporulation (%) | 1 viable spore (%) | 2 viable spores (%) | 3 viable spores (%) | 4 viable spores (%) |
Homozygous normal | 16 | 70±5.81 | 6 | 0 | 38 | 56 |
Heterozygous | 14 | 66.3±4.51 | 0 | 0 | 36 | 64 |
Homozygous mutant | 31 | 52.1±4.11 | 23 | 65 | 13 | 0 |
Mitotic Phenotype of blm1-1 Mutant Strains: Reduced Growth Rate and Elevated Frequencies of Irregularly-shaped Cells
Strains | Final Titers (cells/ml) | Number and Percent of Irregularly- shaped Cells 2 | Normal-sized Single Cells and Budded Cells 3 | Number and Percent of Enlarged Cells 4 | Total Number of Cells |
Means of experiments one and two | |||||
BLM1 (CM1457-79A) | 5.00 x 108 | 0% | (462) 84.6% | (84) 15.4% | 546 (100%) |
Experiments three and four: means of both strains | |||||
blm1-1 (CM1457-79C and CM1457-79D) | 3.60 x 108 | (28.5) 5.23% | (350) 64.1% | (167) 30.6% | 546 (100%) |
Experiments five and six: means of both strains | |||||
blm1-1 (CM1457-79C and CM1457-79D) | 3.78 x 108 | (29) 5.31% | (347) 63.6% | (170) 31.1% | 546 (100%) |
Isolation of Plasmids, Restriction Analyses, and DNA Sequence Analyses
Relief of Drug Hypersensitivity
Genetic Mapping Showed that MSH4 is not the BLM1 Structural Gene
Classes of tetrads | Number of tetrads | Number of spores | blm1-1, leu2 | BLM1, LEU2 | BLM1, leu2 | blm1-1, LEU2 |
Parental Ditype (PD) | 5 | 20 | 10 | 10 | 0 | 0 |
Non-Parental Ditype (NPD) | 6 | 24 | 0 | 0 | 12 | 12 |
Tetratype (TT) | 3 | 12 | 3 | 3 | 3 | 3 |
Total spores | 14 | 56 | 13 | 13 | 15 | 15 |
Discussion
Acknowledgements
References
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Anyatonwu, G.; Garcia, E.; Pramanik, A.; Powell, M.; Moore, C.W. Meiotic and Mitotic Phenotypes Conferred by the blm1-1 Mutation in Saccharomyces cerevisiae and MSH4 Suppression of the Bleomycin Hypersusceptibility. Int. J. Mol. Sci. 2003, 4, 1-12. https://doi.org/10.3390/i4010001
Anyatonwu G, Garcia E, Pramanik A, Powell M, Moore CW. Meiotic and Mitotic Phenotypes Conferred by the blm1-1 Mutation in Saccharomyces cerevisiae and MSH4 Suppression of the Bleomycin Hypersusceptibility. International Journal of Molecular Sciences. 2003; 4(1):1-12. https://doi.org/10.3390/i4010001
Chicago/Turabian StyleAnyatonwu, Georgia, Ediberto Garcia, Ajay Pramanik, Marie Powell, and Carol Wood Moore. 2003. "Meiotic and Mitotic Phenotypes Conferred by the blm1-1 Mutation in Saccharomyces cerevisiae and MSH4 Suppression of the Bleomycin Hypersusceptibility" International Journal of Molecular Sciences 4, no. 1: 1-12. https://doi.org/10.3390/i4010001