{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"omics_type":["Metabolomics","Unknown","Transcriptomics","Genomics","Proteomics"],"submitter":["DCC modENCODE"],"study_type":["ChIP-chip by tiling array"],"organism":["Drosophila melanogaster"],"species":["Drosophila melanogaster"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-GEOD-27803"],"description":["modENCODE_submission_3041 This submission comes from a modENCODE project of Gary Karpen. For full list of modENCODE projects, see http://www.genome.gov/26524648 Project Goal: We aim to determine the locations of the major histone modifications across the Drosophila melanogaster genome. The modifications under study are involved in basic chromosomal functions such as DNA replication, gene expression, gene silencing, and inheritance. We will perform Chromatin ImmunoPrecipitation (ChIP) using genomic tiling arrays. We will initially assay localizations using chromatin from three cell lines and two embryonic stages, and will then extend the analysis of a subset of proteins to four additional animal tissues/stages. For data usage terms and conditions, please refer to http://www.genome.gov/27528"],"repository":["biostudies-arrayexpress"],"sample_protocol":["Nucleic Acid Extraction - 1. Add 30ml of PAS (50% suspension in RIPA (-PMSF)) to 500ml of crosslinked chromatin. Incubate 1h at +40C.   2. Spin suspensions for 2min at top speed +40C. Transfer supernatants to new tubes. Add 5ml of 100mM PMSF solution in isopropanol to each 500ml aliquot of precleared chromatin.   3. Add appropriate amount of antibody to each reaction. Do not forget to set up no Ab control. Incubate for 15 hours at +40C on rotating weel.   4. Add 40ml of PAS (50% suspension in RIPA (-PMSF)), incubate 3h at +40C on rotating weel.   5. Wash the beads 5 times 10min each with 1ml of RIPA, then one time with 1ml of LiCl ChIP buffer and finally twice with 1ml of TE. To pellet the beads between washes spin samples for 20sec +40C at top speed. Do all the washes at +40C.   6. Resusp","Nucleic Acid Extraction - 1. Grow cell cultures in large T-flasks (175-225cm2) to a density of~5x106 cells/ml. Combine the content of 2-3 flasks in one. Immediately proceed with the following steps.   2. Add 37% formaldehyde (Sigma) to final concentration 1.8% directly to the culture incubate for 10min at 250C on a rotating or rocking platform.   3. Stop reaction by adding 1.25M glycine pH 7.0 to each culture to the final concentration of 125mM. Mix well, within next 5 min aliquot cell suspension into 50ml falcon tubes and accumulate them on ice.   4. Pellet fixed cells by spinning for 2? at 1500g, +40C. Use swinging bucket rotor.   5. Resuspend each pellet in 5ml of sterile 1xPBS, pool suspensions together.   6. Pellet the cells by spinning for 2? at 1500g, +40C.   7. Resuspend the cells ","Labeling - 1. Prehybridize array for 1 hr in 200ml of 1xMES-Triton at 450C with 45 rpm rotation.   2. Incubate hybridization cocktail for 10min at 1000C then 10min at 450C. Spin at max speed for 3 min at RT. Transfer the supernatant to new tube. Spin for 3 more minutes use 200ml of resulted supernatant for hybridization.   3. Hybridize 18hr at 450C with 45 rpm rotation.   4. Use fluidics station EukGE-WS2v4 protocol (Affymetrix) for washing and staining.","Nucleic Acid Extraction - After treatment with formaldehyde, Drosophila tissue culture cells were lysed with SDS and sonication (on ice) in the presence of protease inhibitors.  The resulting chromatin was treated with non-ionic detergents at physiological concentrations of monovalent cations.  Ribonucleic acid was removed by RNAse A, and the DNA was purified using SDS, proteinase K, and organic extraction, followed by ethanol precipitation.","Hybridization - 1. Prehybridize array for 1 hr in 200ml of 1xMES-Triton at 450C with 45 rpm rotation.   2. Incubate hybridization cocktail for 10min at 1000C then 10min at 450C. Spin at max speed for 3 min at RT. Transfer the supernatant to new tube. Spin for 3 more minutes use 200ml of resulted supernatant for hybridization.   3. Hybridize 18hr at 450C with 45 rpm rotation.   4. Use fluidics station EukGE-WS2v4 protocol (Affymetrix) for washing and staining."],"figure_sub":["MIAME Score","Raw Data","Organization","Assays and Data","MAGE-TAB Files","Array Designs"],"pubmed_authors":["Nicole Riddle","Gary Karpen","DCC modENCODE","Vincenzo Pirrotta","Andrey Gortchakov","Art Alekseyenko","Greg Shanower","Michael Tolstorukov","Sarah Elgin","Aki Minoda","Mitzi Kuroda","Cameron Kennedy","Peter Park","Yuri Schwartz","Peter Kharchenko"],"data_protocol":["Feature Extraction - M-value normalization protocol. The log-intensity ratio values (M-values) are calculated for all perfect match (PM) probes as log2(ChIP intensity) - log2(input intensity). The M values are then shifted so that the mean is equal to 0.","Image Adquisition - Standard Affymetrix Array Scanning Protocol was used"],"additional_accession":[]},"is_claimable":false,"name":"MOF_Q4145.BG3","description":"modENCODE_submission_3041 This submission comes from a modENCODE project of Gary Karpen. For full list of modENCODE projects, see http://www.genome.gov/26524648 Project Goal: We aim to determine the locations of the major histone modifications across the Drosophila melanogaster genome. The modifications under study are involved in basic chromosomal functions such as DNA replication, gene expression, gene silencing, and inheritance. We will perform Chromatin ImmunoPrecipitation (ChIP) using genomic tiling arrays. We will initially assay localizations using chromatin from three cell lines and two embryonic stages, and will then extend the analysis of a subset of proteins to four additional animal tissues/stages. For data usage terms and conditions, please refer to http://www.genome.gov/27528","dates":{"release":"2011-03-09T00:00:00Z","modification":"2023-08-12T13:03:52.532Z","creation":"2022-02-02T15:09:14.423Z"},"accession":"E-GEOD-27803","cross_references":{"GEO":["GSE27803"],"EFO":["EFO_0002762"]}}