Deletion of Budding Yeast <i<MAD2</i< Suppresses Clone-to-Clone Differences in Artificial Linear Chromosome Copy Numbers and Gives Rise to Higher Retention Rates
Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average arti...
Ausführliche Beschreibung
Gespeichert in:
| Autor*in: |
Scott C. Schuyler [verfasserIn] Lin-Ing Wang [verfasserIn] Yi-Shan Ding [verfasserIn] Yi-Chieh Lee [verfasserIn] Hsin-Yu Chen [verfasserIn] |
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| Format: |
E-Artikel |
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| Sprache: |
Englisch |
| Erschienen: |
2020 |
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| Schlagwörter: |
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| Übergeordnetes Werk: |
In: Microorganisms - MDPI AG, 2013, 8(2020), 10, p 1495 |
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| Übergeordnetes Werk: |
volume:8 ; year:2020 ; number:10, p 1495 |
| Links: |
|---|
| DOI / URN: |
10.3390/microorganisms8101495 |
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| Katalog-ID: |
DOAJ064349365 |
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| 245 | 1 | 0 | |a Deletion of Budding Yeast <i<MAD2</i< Suppresses Clone-to-Clone Differences in Artificial Linear Chromosome Copy Numbers and Gives Rise to Higher Retention Rates |
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| 520 | |a Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average artificial linear chromosome copy numbers in a population of cells, as measured by quantitative polymerase chain reactions (qPCR), and retention rates, as measured by fluctuation analyses, were performed on a total of 62 individual wild type and <i<mad2∆</i< mutant haploid and diploid clones. Wild type cells, both haploids and diploids, displayed phenotypically unique clone-to-clone differences: one group of 15 clones displayed low-copy numbers per cell and high retention rates, were 1 clone was found to have undergone a genomic integration event, and the second group of 15 clones displayed high copy numbers per cell and low retention rates, with the latter values being consistent with the previously published results where only a single clone had been measured. These chromosome states were observed to be unstable when propagated for 10 days under selection, where high copy-low retention rate clones evolved into low copy-high retention rate clones, but no evidence for integration events was observed. By contrast, <i<mad2∆</i< haploid and <i<mad2∆/mad2∆</i< diploids displayed a suppression of the clone-to-clone differences, where 20 out of 21 clones had mid-level artificial linear chromosome copy numbers per cell, but maintained elevated chromosome retention rates. The elevated levels in retention rates in <i<mad2∆</i< and <i<mad2∆</i</<i<mad2∆</i< cells were also maintained even in the absence of selection during growth over 3 days. <i<MAD2/mad2∆</i< heterozygous diploids displayed multiple clonal groups: 4 with low copy numbers, 5 with mid-level copy numbers, and 1 with a high copy number of artificial linear chromosomes, but all 10 clones uniformly displayed low retention rates. Our observations reveal that <i<MAD2</i< function contributes to the ability of yeast cells to maintain a high number of artificial linear chromosomes per cell in some clones, but, counter-intuitively, <i<mad2∆</i< suppresses clone-to-clone differences and leads to an improvement in artificial linear chromosome retention rates yielding a more uniform and stable clonal population with mid-level chromosome copy numbers per cell. | ||
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10.3390/microorganisms8101495 doi (DE-627)DOAJ064349365 (DE-599)DOAJee60080b85f847b59d0b1cbe51b435e8 DE-627 ger DE-627 rakwb eng QH301-705.5 Scott C. Schuyler verfasserin aut Deletion of Budding Yeast <i<MAD2</i< Suppresses Clone-to-Clone Differences in Artificial Linear Chromosome Copy Numbers and Gives Rise to Higher Retention Rates 2020 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average artificial linear chromosome copy numbers in a population of cells, as measured by quantitative polymerase chain reactions (qPCR), and retention rates, as measured by fluctuation analyses, were performed on a total of 62 individual wild type and <i<mad2∆</i< mutant haploid and diploid clones. Wild type cells, both haploids and diploids, displayed phenotypically unique clone-to-clone differences: one group of 15 clones displayed low-copy numbers per cell and high retention rates, were 1 clone was found to have undergone a genomic integration event, and the second group of 15 clones displayed high copy numbers per cell and low retention rates, with the latter values being consistent with the previously published results where only a single clone had been measured. These chromosome states were observed to be unstable when propagated for 10 days under selection, where high copy-low retention rate clones evolved into low copy-high retention rate clones, but no evidence for integration events was observed. By contrast, <i<mad2∆</i< haploid and <i<mad2∆/mad2∆</i< diploids displayed a suppression of the clone-to-clone differences, where 20 out of 21 clones had mid-level artificial linear chromosome copy numbers per cell, but maintained elevated chromosome retention rates. The elevated levels in retention rates in <i<mad2∆</i< and <i<mad2∆</i</<i<mad2∆</i< cells were also maintained even in the absence of selection during growth over 3 days. <i<MAD2/mad2∆</i< heterozygous diploids displayed multiple clonal groups: 4 with low copy numbers, 5 with mid-level copy numbers, and 1 with a high copy number of artificial linear chromosomes, but all 10 clones uniformly displayed low retention rates. Our observations reveal that <i<MAD2</i< function contributes to the ability of yeast cells to maintain a high number of artificial linear chromosomes per cell in some clones, but, counter-intuitively, <i<mad2∆</i< suppresses clone-to-clone differences and leads to an improvement in artificial linear chromosome retention rates yielding a more uniform and stable clonal population with mid-level chromosome copy numbers per cell. budding yeast <i<Saccharomyces cerevisiae</i< chromosome mitotic spindle checkpoint Mitotic Arrest-Deficient 2 (<i<MAD2</i<) quantitative polymerase chain reaction (qPCR) Biology (General) Lin-Ing Wang verfasserin aut Yi-Shan Ding verfasserin aut Yi-Chieh Lee verfasserin aut Hsin-Yu Chen verfasserin aut In Microorganisms MDPI AG, 2013 8(2020), 10, p 1495 (DE-627)750370696 (DE-600)2720891-6 20762607 nnns volume:8 year:2020 number:10, p 1495 https://doi.org/10.3390/microorganisms8101495 kostenfrei https://doaj.org/article/ee60080b85f847b59d0b1cbe51b435e8 kostenfrei https://www.mdpi.com/2076-2607/8/10/1495 kostenfrei https://doaj.org/toc/2076-2607 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 8 2020 10, p 1495 |
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10.3390/microorganisms8101495 doi (DE-627)DOAJ064349365 (DE-599)DOAJee60080b85f847b59d0b1cbe51b435e8 DE-627 ger DE-627 rakwb eng QH301-705.5 Scott C. Schuyler verfasserin aut Deletion of Budding Yeast <i<MAD2</i< Suppresses Clone-to-Clone Differences in Artificial Linear Chromosome Copy Numbers and Gives Rise to Higher Retention Rates 2020 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average artificial linear chromosome copy numbers in a population of cells, as measured by quantitative polymerase chain reactions (qPCR), and retention rates, as measured by fluctuation analyses, were performed on a total of 62 individual wild type and <i<mad2∆</i< mutant haploid and diploid clones. Wild type cells, both haploids and diploids, displayed phenotypically unique clone-to-clone differences: one group of 15 clones displayed low-copy numbers per cell and high retention rates, were 1 clone was found to have undergone a genomic integration event, and the second group of 15 clones displayed high copy numbers per cell and low retention rates, with the latter values being consistent with the previously published results where only a single clone had been measured. These chromosome states were observed to be unstable when propagated for 10 days under selection, where high copy-low retention rate clones evolved into low copy-high retention rate clones, but no evidence for integration events was observed. By contrast, <i<mad2∆</i< haploid and <i<mad2∆/mad2∆</i< diploids displayed a suppression of the clone-to-clone differences, where 20 out of 21 clones had mid-level artificial linear chromosome copy numbers per cell, but maintained elevated chromosome retention rates. The elevated levels in retention rates in <i<mad2∆</i< and <i<mad2∆</i</<i<mad2∆</i< cells were also maintained even in the absence of selection during growth over 3 days. <i<MAD2/mad2∆</i< heterozygous diploids displayed multiple clonal groups: 4 with low copy numbers, 5 with mid-level copy numbers, and 1 with a high copy number of artificial linear chromosomes, but all 10 clones uniformly displayed low retention rates. Our observations reveal that <i<MAD2</i< function contributes to the ability of yeast cells to maintain a high number of artificial linear chromosomes per cell in some clones, but, counter-intuitively, <i<mad2∆</i< suppresses clone-to-clone differences and leads to an improvement in artificial linear chromosome retention rates yielding a more uniform and stable clonal population with mid-level chromosome copy numbers per cell. budding yeast <i<Saccharomyces cerevisiae</i< chromosome mitotic spindle checkpoint Mitotic Arrest-Deficient 2 (<i<MAD2</i<) quantitative polymerase chain reaction (qPCR) Biology (General) Lin-Ing Wang verfasserin aut Yi-Shan Ding verfasserin aut Yi-Chieh Lee verfasserin aut Hsin-Yu Chen verfasserin aut In Microorganisms MDPI AG, 2013 8(2020), 10, p 1495 (DE-627)750370696 (DE-600)2720891-6 20762607 nnns volume:8 year:2020 number:10, p 1495 https://doi.org/10.3390/microorganisms8101495 kostenfrei https://doaj.org/article/ee60080b85f847b59d0b1cbe51b435e8 kostenfrei https://www.mdpi.com/2076-2607/8/10/1495 kostenfrei https://doaj.org/toc/2076-2607 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 8 2020 10, p 1495 |
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10.3390/microorganisms8101495 doi (DE-627)DOAJ064349365 (DE-599)DOAJee60080b85f847b59d0b1cbe51b435e8 DE-627 ger DE-627 rakwb eng QH301-705.5 Scott C. Schuyler verfasserin aut Deletion of Budding Yeast <i<MAD2</i< Suppresses Clone-to-Clone Differences in Artificial Linear Chromosome Copy Numbers and Gives Rise to Higher Retention Rates 2020 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average artificial linear chromosome copy numbers in a population of cells, as measured by quantitative polymerase chain reactions (qPCR), and retention rates, as measured by fluctuation analyses, were performed on a total of 62 individual wild type and <i<mad2∆</i< mutant haploid and diploid clones. Wild type cells, both haploids and diploids, displayed phenotypically unique clone-to-clone differences: one group of 15 clones displayed low-copy numbers per cell and high retention rates, were 1 clone was found to have undergone a genomic integration event, and the second group of 15 clones displayed high copy numbers per cell and low retention rates, with the latter values being consistent with the previously published results where only a single clone had been measured. These chromosome states were observed to be unstable when propagated for 10 days under selection, where high copy-low retention rate clones evolved into low copy-high retention rate clones, but no evidence for integration events was observed. By contrast, <i<mad2∆</i< haploid and <i<mad2∆/mad2∆</i< diploids displayed a suppression of the clone-to-clone differences, where 20 out of 21 clones had mid-level artificial linear chromosome copy numbers per cell, but maintained elevated chromosome retention rates. The elevated levels in retention rates in <i<mad2∆</i< and <i<mad2∆</i</<i<mad2∆</i< cells were also maintained even in the absence of selection during growth over 3 days. <i<MAD2/mad2∆</i< heterozygous diploids displayed multiple clonal groups: 4 with low copy numbers, 5 with mid-level copy numbers, and 1 with a high copy number of artificial linear chromosomes, but all 10 clones uniformly displayed low retention rates. Our observations reveal that <i<MAD2</i< function contributes to the ability of yeast cells to maintain a high number of artificial linear chromosomes per cell in some clones, but, counter-intuitively, <i<mad2∆</i< suppresses clone-to-clone differences and leads to an improvement in artificial linear chromosome retention rates yielding a more uniform and stable clonal population with mid-level chromosome copy numbers per cell. budding yeast <i<Saccharomyces cerevisiae</i< chromosome mitotic spindle checkpoint Mitotic Arrest-Deficient 2 (<i<MAD2</i<) quantitative polymerase chain reaction (qPCR) Biology (General) Lin-Ing Wang verfasserin aut Yi-Shan Ding verfasserin aut Yi-Chieh Lee verfasserin aut Hsin-Yu Chen verfasserin aut In Microorganisms MDPI AG, 2013 8(2020), 10, p 1495 (DE-627)750370696 (DE-600)2720891-6 20762607 nnns volume:8 year:2020 number:10, p 1495 https://doi.org/10.3390/microorganisms8101495 kostenfrei https://doaj.org/article/ee60080b85f847b59d0b1cbe51b435e8 kostenfrei https://www.mdpi.com/2076-2607/8/10/1495 kostenfrei https://doaj.org/toc/2076-2607 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 8 2020 10, p 1495 |
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10.3390/microorganisms8101495 doi (DE-627)DOAJ064349365 (DE-599)DOAJee60080b85f847b59d0b1cbe51b435e8 DE-627 ger DE-627 rakwb eng QH301-705.5 Scott C. Schuyler verfasserin aut Deletion of Budding Yeast <i<MAD2</i< Suppresses Clone-to-Clone Differences in Artificial Linear Chromosome Copy Numbers and Gives Rise to Higher Retention Rates 2020 Text txt rdacontent Computermedien c rdamedia Online-Ressource cr rdacarrier Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average artificial linear chromosome copy numbers in a population of cells, as measured by quantitative polymerase chain reactions (qPCR), and retention rates, as measured by fluctuation analyses, were performed on a total of 62 individual wild type and <i<mad2∆</i< mutant haploid and diploid clones. Wild type cells, both haploids and diploids, displayed phenotypically unique clone-to-clone differences: one group of 15 clones displayed low-copy numbers per cell and high retention rates, were 1 clone was found to have undergone a genomic integration event, and the second group of 15 clones displayed high copy numbers per cell and low retention rates, with the latter values being consistent with the previously published results where only a single clone had been measured. These chromosome states were observed to be unstable when propagated for 10 days under selection, where high copy-low retention rate clones evolved into low copy-high retention rate clones, but no evidence for integration events was observed. By contrast, <i<mad2∆</i< haploid and <i<mad2∆/mad2∆</i< diploids displayed a suppression of the clone-to-clone differences, where 20 out of 21 clones had mid-level artificial linear chromosome copy numbers per cell, but maintained elevated chromosome retention rates. The elevated levels in retention rates in <i<mad2∆</i< and <i<mad2∆</i</<i<mad2∆</i< cells were also maintained even in the absence of selection during growth over 3 days. <i<MAD2/mad2∆</i< heterozygous diploids displayed multiple clonal groups: 4 with low copy numbers, 5 with mid-level copy numbers, and 1 with a high copy number of artificial linear chromosomes, but all 10 clones uniformly displayed low retention rates. Our observations reveal that <i<MAD2</i< function contributes to the ability of yeast cells to maintain a high number of artificial linear chromosomes per cell in some clones, but, counter-intuitively, <i<mad2∆</i< suppresses clone-to-clone differences and leads to an improvement in artificial linear chromosome retention rates yielding a more uniform and stable clonal population with mid-level chromosome copy numbers per cell. budding yeast <i<Saccharomyces cerevisiae</i< chromosome mitotic spindle checkpoint Mitotic Arrest-Deficient 2 (<i<MAD2</i<) quantitative polymerase chain reaction (qPCR) Biology (General) Lin-Ing Wang verfasserin aut Yi-Shan Ding verfasserin aut Yi-Chieh Lee verfasserin aut Hsin-Yu Chen verfasserin aut In Microorganisms MDPI AG, 2013 8(2020), 10, p 1495 (DE-627)750370696 (DE-600)2720891-6 20762607 nnns volume:8 year:2020 number:10, p 1495 https://doi.org/10.3390/microorganisms8101495 kostenfrei https://doaj.org/article/ee60080b85f847b59d0b1cbe51b435e8 kostenfrei https://www.mdpi.com/2076-2607/8/10/1495 kostenfrei https://doaj.org/toc/2076-2607 Journal toc kostenfrei GBV_USEFLAG_A SYSFLAG_A GBV_DOAJ GBV_ILN_20 GBV_ILN_22 GBV_ILN_23 GBV_ILN_24 GBV_ILN_39 GBV_ILN_40 GBV_ILN_62 GBV_ILN_63 GBV_ILN_65 GBV_ILN_69 GBV_ILN_70 GBV_ILN_73 GBV_ILN_74 GBV_ILN_95 GBV_ILN_105 GBV_ILN_110 GBV_ILN_151 GBV_ILN_161 GBV_ILN_170 GBV_ILN_213 GBV_ILN_230 GBV_ILN_285 GBV_ILN_293 GBV_ILN_602 GBV_ILN_2014 GBV_ILN_4012 GBV_ILN_4037 GBV_ILN_4112 GBV_ILN_4125 GBV_ILN_4126 GBV_ILN_4249 GBV_ILN_4305 GBV_ILN_4306 GBV_ILN_4307 GBV_ILN_4313 GBV_ILN_4322 GBV_ILN_4323 GBV_ILN_4324 GBV_ILN_4325 GBV_ILN_4338 GBV_ILN_4367 GBV_ILN_4700 AR 8 2020 10, p 1495 |
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Deletion of Budding Yeast <i<MAD2</i< Suppresses Clone-to-Clone Differences in Artificial Linear Chromosome Copy Numbers and Gives Rise to Higher Retention Rates |
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Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average artificial linear chromosome copy numbers in a population of cells, as measured by quantitative polymerase chain reactions (qPCR), and retention rates, as measured by fluctuation analyses, were performed on a total of 62 individual wild type and <i<mad2∆</i< mutant haploid and diploid clones. Wild type cells, both haploids and diploids, displayed phenotypically unique clone-to-clone differences: one group of 15 clones displayed low-copy numbers per cell and high retention rates, were 1 clone was found to have undergone a genomic integration event, and the second group of 15 clones displayed high copy numbers per cell and low retention rates, with the latter values being consistent with the previously published results where only a single clone had been measured. These chromosome states were observed to be unstable when propagated for 10 days under selection, where high copy-low retention rate clones evolved into low copy-high retention rate clones, but no evidence for integration events was observed. By contrast, <i<mad2∆</i< haploid and <i<mad2∆/mad2∆</i< diploids displayed a suppression of the clone-to-clone differences, where 20 out of 21 clones had mid-level artificial linear chromosome copy numbers per cell, but maintained elevated chromosome retention rates. The elevated levels in retention rates in <i<mad2∆</i< and <i<mad2∆</i</<i<mad2∆</i< cells were also maintained even in the absence of selection during growth over 3 days. <i<MAD2/mad2∆</i< heterozygous diploids displayed multiple clonal groups: 4 with low copy numbers, 5 with mid-level copy numbers, and 1 with a high copy number of artificial linear chromosomes, but all 10 clones uniformly displayed low retention rates. Our observations reveal that <i<MAD2</i< function contributes to the ability of yeast cells to maintain a high number of artificial linear chromosomes per cell in some clones, but, counter-intuitively, <i<mad2∆</i< suppresses clone-to-clone differences and leads to an improvement in artificial linear chromosome retention rates yielding a more uniform and stable clonal population with mid-level chromosome copy numbers per cell. |
| abstractGer |
Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average artificial linear chromosome copy numbers in a population of cells, as measured by quantitative polymerase chain reactions (qPCR), and retention rates, as measured by fluctuation analyses, were performed on a total of 62 individual wild type and <i<mad2∆</i< mutant haploid and diploid clones. Wild type cells, both haploids and diploids, displayed phenotypically unique clone-to-clone differences: one group of 15 clones displayed low-copy numbers per cell and high retention rates, were 1 clone was found to have undergone a genomic integration event, and the second group of 15 clones displayed high copy numbers per cell and low retention rates, with the latter values being consistent with the previously published results where only a single clone had been measured. These chromosome states were observed to be unstable when propagated for 10 days under selection, where high copy-low retention rate clones evolved into low copy-high retention rate clones, but no evidence for integration events was observed. By contrast, <i<mad2∆</i< haploid and <i<mad2∆/mad2∆</i< diploids displayed a suppression of the clone-to-clone differences, where 20 out of 21 clones had mid-level artificial linear chromosome copy numbers per cell, but maintained elevated chromosome retention rates. The elevated levels in retention rates in <i<mad2∆</i< and <i<mad2∆</i</<i<mad2∆</i< cells were also maintained even in the absence of selection during growth over 3 days. <i<MAD2/mad2∆</i< heterozygous diploids displayed multiple clonal groups: 4 with low copy numbers, 5 with mid-level copy numbers, and 1 with a high copy number of artificial linear chromosomes, but all 10 clones uniformly displayed low retention rates. Our observations reveal that <i<MAD2</i< function contributes to the ability of yeast cells to maintain a high number of artificial linear chromosomes per cell in some clones, but, counter-intuitively, <i<mad2∆</i< suppresses clone-to-clone differences and leads to an improvement in artificial linear chromosome retention rates yielding a more uniform and stable clonal population with mid-level chromosome copy numbers per cell. |
| abstract_unstemmed |
Our goal was to investigate the changes in artificial short-linear chromosome average copy numbers per cell arising from partial or full loss of Mitotic Arrest-Deficient 2 (<i<MAD2</i<) spindle checkpoint function in budding yeast <i<Saccharomyces cerevisiae</i<. Average artificial linear chromosome copy numbers in a population of cells, as measured by quantitative polymerase chain reactions (qPCR), and retention rates, as measured by fluctuation analyses, were performed on a total of 62 individual wild type and <i<mad2∆</i< mutant haploid and diploid clones. Wild type cells, both haploids and diploids, displayed phenotypically unique clone-to-clone differences: one group of 15 clones displayed low-copy numbers per cell and high retention rates, were 1 clone was found to have undergone a genomic integration event, and the second group of 15 clones displayed high copy numbers per cell and low retention rates, with the latter values being consistent with the previously published results where only a single clone had been measured. These chromosome states were observed to be unstable when propagated for 10 days under selection, where high copy-low retention rate clones evolved into low copy-high retention rate clones, but no evidence for integration events was observed. By contrast, <i<mad2∆</i< haploid and <i<mad2∆/mad2∆</i< diploids displayed a suppression of the clone-to-clone differences, where 20 out of 21 clones had mid-level artificial linear chromosome copy numbers per cell, but maintained elevated chromosome retention rates. The elevated levels in retention rates in <i<mad2∆</i< and <i<mad2∆</i</<i<mad2∆</i< cells were also maintained even in the absence of selection during growth over 3 days. <i<MAD2/mad2∆</i< heterozygous diploids displayed multiple clonal groups: 4 with low copy numbers, 5 with mid-level copy numbers, and 1 with a high copy number of artificial linear chromosomes, but all 10 clones uniformly displayed low retention rates. Our observations reveal that <i<MAD2</i< function contributes to the ability of yeast cells to maintain a high number of artificial linear chromosomes per cell in some clones, but, counter-intuitively, <i<mad2∆</i< suppresses clone-to-clone differences and leads to an improvement in artificial linear chromosome retention rates yielding a more uniform and stable clonal population with mid-level chromosome copy numbers per cell. |
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| title_short |
Deletion of Budding Yeast <i<MAD2</i< Suppresses Clone-to-Clone Differences in Artificial Linear Chromosome Copy Numbers and Gives Rise to Higher Retention Rates |
| url |
https://doi.org/10.3390/microorganisms8101495 https://doaj.org/article/ee60080b85f847b59d0b1cbe51b435e8 https://www.mdpi.com/2076-2607/8/10/1495 https://doaj.org/toc/2076-2607 |
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Lin-Ing Wang Yi-Shan Ding Yi-Chieh Lee Hsin-Yu Chen |
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