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. 1999 Oct 18;147(2):335–350. doi: 10.1083/jcb.147.2.335

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© 1999 The Rockefeller University Press

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/).

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graphic file with name JCB9907049.f5a.jpg — Bipolar spindle assembly and structural integrity requires either Kar3p or Kip3p. (A) Cells of the indicated genotypes also expressed the SPB component Nuf2p tagged with GFP. Cultures of the indicated genotypes were synchronized at 26°C with α-factor and released into media at 35°C. After 60 min, samples of live yeast cells were immediately observed under the microscope. Examples of wild-type (top row) and kar3Δ kip3Δ (pkar3-64) (bottom row) cells. (B) Quantitation of spindle assembly proficiency. Cells released from α-factor at 35°C, as in A, were observed and the percentage of cells exhibiting two clearly separated GFP dots was determined. The decreases observed in the later time point are due to cells that have entered the next cell cycle. (C) Quantitation of ability to maintain spindle structural integrity. Cells of the indicated genotype (and expressing Nuf2p-GFP) were synchronized at 26°C with hydroxyurea and shifted to 35°C for the indicated times, at which point the percentage of cells exhibiting two clearly separated GFP dots was determined. Strains used: wild-type (MAY5776), kar3Δ (pkar3-64) (MAY5777), kar3Δ kip3Δ (pkar3-64) (MAY5778), kar3Δ dyn1Δ (pkar3-64) (MAY5779), and kip3Δ (pkip3-14) (MAY6030).

graphic file with name JCB9907049.f5b.jpg — Bipolar spindle assembly and structural integrity requires either Kar3p or Kip3p. (A) Cells of the indicated genotypes also expressed the SPB component Nuf2p tagged with GFP. Cultures of the indicated genotypes were synchronized at 26°C with α-factor and released into media at 35°C. After 60 min, samples of live yeast cells were immediately observed under the microscope. Examples of wild-type (top row) and kar3Δ kip3Δ (pkar3-64) (bottom row) cells. (B) Quantitation of spindle assembly proficiency. Cells released from α-factor at 35°C, as in A, were observed and the percentage of cells exhibiting two clearly separated GFP dots was determined. The decreases observed in the later time point are due to cells that have entered the next cell cycle. (C) Quantitation of ability to maintain spindle structural integrity. Cells of the indicated genotype (and expressing Nuf2p-GFP) were synchronized at 26°C with hydroxyurea and shifted to 35°C for the indicated times, at which point the percentage of cells exhibiting two clearly separated GFP dots was determined. Strains used: wild-type (MAY5776), kar3Δ (pkar3-64) (MAY5777), kar3Δ kip3Δ (pkar3-64) (MAY5778), kar3Δ dyn1Δ (pkar3-64) (MAY5779), and kip3Δ (pkip3-14) (MAY6030).