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Analysis of a Double-Strand DNA Break in living S. cerevisiae Cells

Analysis of a Double-Strand DNA Break in living S. cerevisiae Cells. Leana M. Topper Kerry S. Bloom. Department of Biology, University of North Carolina at Chapel Hill;. Chromosome III. MAT. CEN. ~85 kb. LacO array. ~500 bp. MAT. LacO. KpnI. KpnI. probe. 2.5 kb.

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Analysis of a Double-Strand DNA Break in living S. cerevisiae Cells

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  1. Analysis of a Double-Strand DNA Break in living S. cerevisiae Cells Leana M. Topper Kerry S. Bloom Department of Biology, University of North Carolina at Chapel Hill;

  2. Chromosome III MAT CEN ~85 kb LacO array ~500 bp

  3. MAT LacO KpnI KpnI probe 2.5 kb No pGalHOT W/ pGalHOT 0 0.5 2 3 4 1 0 0.5 2 3 4 1 h post gal 8 kb KpnI fragment HO cut fragment

  4. 6:00 7:00 2:00 10:00 13:00 15:00 8:00 Live cell after HO induction

  5. Movement of Spindle Pole Bodies and lacO in live cells following HO expression Average film time: 13 min No bud: 13 Small-budded cells: 8 Large-budded cells: 29 Average spindle length = 1.69 ±0.32mm Range = 1.05-2.44 mm

  6. Population analysis of lacO and SPBs after induction of HO

  7. Deletion of Rad52 does not affect LacO movement 6:00 2:00 0:00 9:00 21:00 15:00 20:00

  8. Formation of Rad52 foci following DNA damage

  9. Formation of Rad52-GFP foci after induction of HO

  10. Rad52-GFP foci and Spindle Pole Bodies Move Independently 7:00 5:00 1:00 17:00 18:00 8:00 12:00

  11. Rad52-CFP and LacO spots do not colocalize 6:00 8:00 3:00 16:00 9:00 11:00 14:00

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