Comment[ArrayExpressAccession] E-GEOD-50988 MAGE-TAB Version 1.1 Public Release Date 2013-10-11 Investigation Title Identification of cell cycle-regulated genes periodically expressed in U2OS cells and their regulation by FOXM1 and E2F transcription factors Comment[Submitted Name] Identification of cell cycle-regulated genes periodically expressed in U2OS cells and their regulation by FOXM1 and E2F transcription factors Experiment Description We identified the cell cycle-regulated mRNA transcripts genome-wide in the osteosarcoma derived U2OS cell line. This resulted in 2,140 transcripts mapping to 1,871 unique cell cycle-regulated genes that show periodic oscillations across multiple synchronous cell cycles. We identified genomic loci bound by the G2/M transcription factor FOXM1 by Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) and associated these with cell cycle-regulated genes. FOXM1 was bound to cell cycle-regulated genes with peak expression in both S phase and G2/M phases. ChIP-seq genomic loci were shown to be responsive to FOXM1 using a real-time luciferase assay in live cells, showing that FOXM1 strongly activates promoters of G2/M phase genes and weakly activates those induced in S phase. Analysis of ChIP-seq data from a panel of cell cycle-transcription factors (E2F1, E2F4, E2F6, and GABPA) from ENCODE and ChIP-seq data for the DREAM complex, found that a set of core cell cycle genes regulated in both U2OS and HeLa cells are bound by multiple cell cycle transcription factors. These data identify the cell cycle regulated genes in a second cancer derived cell line and provide a comprehensive picture of the transcriptional regulatory systems controlling periodic gene expression in the human cell division cycle. Cell cycle-regulated gene expression identified from three double thymidine time courses and one thymidine nocodazole time course. Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Grant Grant Whitfield Person First Name Gavin Gavin Michael Person Mid Initials D D L Person Email geo@ncbi.nlm.nih.gov Person Affiliation Geisel School of Medicine Person Address Genetics, Geisel School of Medicine, 7400 Remsen, Hanover, NH, USA Person Roles submitter Protocol Name P-GSE50988-1 P-GSE50988-5 P-GSE50988-7 P-GSE50988-93 P-GSE50988-81 P-GSE50988-65 P-GSE50988-57 P-GSE50988-59 P-GSE50988-56 P-GSE50988-15 P-GSE50988-88 P-GSE50988-64 P-GSE50988-37 P-GSE50988-16 P-GSE50988-23 P-GSE50988-63 P-GSE50988-50 P-GSE50988-40 P-GSE50988-26 P-GSE50988-91 P-GSE50988-14 P-GSE50988-55 P-GSE50988-2 P-GSE50988-60 P-GSE50988-19 P-GSE50988-83 P-GSE50988-43 P-GSE50988-12 P-GSE50988-33 P-GSE50988-38 P-GSE50988-34 P-GSE50988-31 P-GSE50988-49 P-GSE50988-39 P-GSE50988-90 P-GSE50988-17 P-GSE50988-24 P-GSE50988-94 P-GSE50988-78 P-GSE50988-54 P-GSE50988-62 P-GSE50988-18 P-GSE50988-75 P-GSE50988-25 P-GSE50988-70 P-GSE50988-27 P-GSE50988-96 P-GSE50988-76 P-GSE50988-58 P-GSE50988-97 P-GSE50988-11 P-GSE50988-85 P-GSE50988-20 P-GSE50988-98 P-GSE50988-13 P-GSE50988-22 P-GSE50988-51 P-GSE50988-29 P-GSE50988-52 P-GSE50988-80 P-GSE50988-47 P-GSE50988-82 P-GSE50988-9 P-GSE50988-28 P-GSE50988-61 P-GSE50988-86 P-GSE50988-79 P-GSE50988-73 P-GSE50988-46 P-GSE50988-45 P-GSE50988-66 P-GSE50988-36 P-GSE50988-48 P-GSE50988-84 P-GSE50988-92 P-GSE50988-87 P-GSE50988-72 P-GSE50988-21 P-GSE50988-71 P-GSE50988-68 P-GSE50988-95 P-GSE50988-35 P-GSE50988-30 P-GSE50988-67 P-GSE50988-41 P-GSE50988-42 P-GSE50988-6 P-GSE50988-10 P-GSE50988-69 P-GSE50988-77 P-GSE50988-89 P-GSE50988-74 P-GSE50988-32 P-GSE50988-44 P-GSE50988-53 P-GSE50988-3 P-GSE50988-4 P-GSE50988-8 Protocol Description Data for both channels were Lowess-normalized and then the log(2) ratio was taken ID_REF = Spot Reference ID VALUE = log2_ratio of CH2DL_MEAN over CH1DL_MEAN (LOG_RAT2L_MEAN) SPOT = spot number on array CH1_MEAN = CH1_SD = standard deviation of channel 1 intensity CH1_BKD_MEDIAN = channel 1 background median intensity CH1_BKD_SD = standard deviation of channel 1 background median intensity CH2_MEAN = channel 2 mean intensity CH2_SD = standard deviation of channel 2 intensity CH2_BKD_MEDIAN = channel 2 background median intensity CH2_BKD_SD = standard deviation of channel 2 background median intensity TOT_BPIX = number of background pixels TOT_SPIX = number of spot pixels CH2BN_MEDIAN = channel 2 normalized background median intensity CH2IN_MEAN = channel 2 normalized mean intensity CH1DL_MEAN = channel 1 Lowess_normalized mean intensity CH2DL_MEAN = channel 2 Lowess_normalized mean intensity LOG_RAT2N_MEAN = log2_ratio of channel 2 normalized over channel 1 (global normalization) CORR = correlation coefficient among pixels FLAG = Spot flag. 0:not flagged; negative:flagged as bad spots; positive:flagged as good spots Labeled using Agilent Quick Amp Labeling Kit Hyb using Agilent Hyb Kit 34 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 10 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 26 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 10 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 14 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 8 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 14 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 24 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 24 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 10 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 16 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 30 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 22 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 36 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 16 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 36 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 30 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 12 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 6 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. Total RNA isolated using Qiagen Rneasy kit 16 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 22 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 14 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 22 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 8 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 2 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 12 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 4 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 46 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 34 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 14 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 28 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 18 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 32 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 36 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 4 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells and added back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 4 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 20 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 20 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 46 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 34 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 36 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 38 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 40 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells and added back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 0 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells and added back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 12 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 42 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 6 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 18 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cellsand added back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 24 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 44 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cellsand added back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 10 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 28 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 38 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 42 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 0 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 8 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells and added back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 30 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 12 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 2 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 40 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 18 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 20 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 6 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 42 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 28 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 26 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 28 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 8 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 32 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 16 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 32 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 22 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 40 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 26 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 38 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 32 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 38 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 6 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 44 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 30 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 18 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 20 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 0 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 4 hr timepoint. U2OS, double thymidine synch 10-3-07, 2.5mM thymidine. Plated 3*10^5 cells per plate, 18 hr block then 8 hr release, the 18hr block. RNA samples taken every 2hrs. 34 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 2 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells and added back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 26 hr timepoint. U2OS thymidine-nocodazole synch 6-4-08. Plated 3*10^5 cells per p100. grew overnight. 5am added thy to 2.5mM arrested for 17hrs. at 10pm washed cells and added 100nm Noc to cells. 10AM released cells washed. saved floating cells andadded back to plates. Took RNA, protein, and FACS samples (1 plate each) every two hours saving floating cells for first 6 hours. 44 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. 0 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 24 hr timepoint. U2OS, double thymidine synch, 2.5mM thymidine. plated 2.5*10^5 cells per plate, 18hr block then 8hr release, then 18hr block, RNA sample taken every 2 hrs. 2 hr timepoint. U2OS double thymidine synch. 2.5mM thy. Plated 2.7e5 cells per p100 on 10-26-09. 18hr block, 8hr release, 18hr block. RNA samples taken every 2 hrs. DMEM (with 4.5 g/L glucose, L-glu, and sodium pyruvate) + 10% FBS, and 100U penn/strep. Humidified 37C incubator Total RNA isolated using Qiagen Rneasy kit Fluorescent array images were collected for both Cy3 and Cy5 with a GenePix 4000B fluorescent scanner and image intensity data were extracted and analyzed with GenePix Pro analysis software. Protocol Type normalization data transformation protocol labelling protocol hybridization protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol sample treatment protocol growth protocol nucleic acid extraction protocol array scanning protocol Experimental Factor Name CELLS Experimental Factor Type cells Publication Title Identification of cell cycle-regulated genes periodically expressed in U2OS cells and their regulation by FOXM1 and E2F transcription factors. Publication Author List Grant GD, Brooks L 3rd, Zhang X, Mahoney JM, Martyanov V, Wood TA, Sherlock G, Cheng C, Whitfield ML PubMed ID 24109597 Publication DOI 10.1091/mbc.E13-05-0264 Comment[SecondaryAccession] GSE50988 Comment[GEOReleaseDate] 2013-10-11 Comment[ArrayExpressSubmissionDate] 2013-09-18 Comment[GEOLastUpdateDate] 2013-10-12 Comment[AEExperimentType] transcription profiling by array SDRF File E-GEOD-50988.sdrf.txt