Ation on the synaptonemal complex in yeast meiosis. Genes Cells 15: 1036?050. Communicating editor: N. HunterH. Sasanuma et al.GENETICSSupporting Details http:/ /genetics.org/lookup/suppl/doi:10.1534/genetics.113.150615/-/DCRemodeling in the Rad51 DNA Strand-Exchange Protein by the Srs2 HelicaseHiroyuki Sasanuma, Yuko Furihata, Miki Shinohara, and Akira ShinoharaCopyright ?2013 by the Genetics Society of America DOI: 10.1534/genetics.113.Figure S1 Expression of Srs2 in the course of meiosis. (A) SRS2 and NDT80 transcripts through meiosis in wild kind cells (NKY1303/1543) had been analyzed by Northern blotting. Blots for SRS2 and NDT80 in addition to Ethidium bromidestained gel image for loading manage are shown (left). Quantification of signals on blots is shown in suitable. Relative intensities of amounts of SRS2 mRNA at each time point are normalized against that at 0 hr and shown. (B) Meiosis progression of 2HASRS2 strain (HSY560/562) was analyzed by DAPI staining. At the least 150 cells were counted for each and every time point. Meiosis I, open circles; meiosis II, closed circles. (C) Expression of 2HASrs2, Rec8 proteins and tubulin through meiosis of 2HASRS2 cells (HSY560/562) was analyzed by western blotting. Rec8 shows slower migrating species for the duration of meiosis by phosphorylation. An asterisk indicates nonspecific signals.two SIH. Sasanuma et al.(D) Expression of HASrs2 protein through meiosis of your ndt80 (HSY611/613) and ndt80 mnd2 (HSY685/686) mutant cells had been analyzed by western blotting. Rec8 and tubulin were analyzed as a handle. (E) SRS2 and NDT80 transcripts during meiosis in DMC1pSRS2 cells (HSY475/477) had been analyzed by Northern blotting. Blots for SRS2 and NDT80 in addition to Ethidium bromidestained gel image are shown (left). Quantification of signals on blots is shown in suitable. Relative intensities of amounts of SRS2 mRNA at every time point are normalized against that at 0 hr and shown.H. Sasanuma et al.3 SIFigure S2 Overexpression of Srs2 reduces spore viabilities.Buy5-Oxaspiro[2.4]heptane-1-carboxylic acid Distribution of quantity of viable spores per a tetrad is shown for wild kind (left; NKY1303/1543), DMC1pSRS2 heterozygous (middle; NKY1303/HSY477) and homozygous (ideal; HSY474/477) cells. 96 tetrads had been dissected for every single strain.four SIH. Sasanuma et al.Figure S3 Overexpression of Srs2 throughout meiosis. (A, B) SRS2 transcripts for the duration of meiosis in mei5 GALpSRS2 cells (HSY781/783) after the addition of estradiol at 5 hours of meiosis were analyzed by Northern blotting. Blots for SRS2 along with Ethidium bromidestained gel image are shown (A). Quantification of SRS2 signals on blots is shown in (B). Relative intensities of amounts of SRS2 mRNA at every single time point are normalized against that at 0 hr and shown.Ethyl 2-amino-1H-imidazole-5-carboxylate web (C) Induction of Srs2 protein in several strains was verified by western blotting.PMID:24406011 As a manage, an quantity of tubulin can also be studied. Wild variety, NKY1303/1543; srs2, HSY310/315; GALpSRS2, HSY781/783; GALpsrs2K41A, HSY1086/1088; GALpsrs2 (875902); HSY1064/1065. (D) Relative amounts of wild sort Srs2 and tubulin have been studies by western blotting just after serial dilutions. Twofolds cell lysates from wild sort cells (NKY1303/1543) at 4 hr (left lane) relative these from GALpSRS2 (HSY781/783) cells at 8 hr (3 hr induction, correct lane) had been analyzed.H. Sasanuma et al.5 SITable S1 Strain list Name NKY1303 NKY1543 HSY475 HSY477 HSY560 HSY562 HSY310 HSY315 HSY516 HSY520 Genotype MATa ho::LYS2 ura3 leu2::hisG lys2 his4B-LEU2, arg4-bgl MAT ho::LYS2 ura3 leu2::hisG, ly.