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Sample GSM927197 Query DataSets for GSM927197
Status Public on May 05, 2012
Title CG42670_RIP-Seq, rep1
Sample type SRA
 
Source name Drosophila tissue culture cells, CG42670
Organism Drosophila melanogaster
Characteristics cell line: S2R+
orf encoding protein: CG42670
rip antibody: HA Immunoaffinity Resin (Sigma)
Treatment protocol Open reading frames for the indicated proteins were inserted into pMK33-C-FLAG-HA acceptor vector. Each clone was transiently transfected into a culture of Drosophila S2R+ cells. Protein expression was induced with 0.35 mM CuSO4.
Growth protocol Cells were cultured in Schneider's medium at 25˚C.
Extracted molecule total RNA
Extraction protocol Whole-cell lysates were prepared in lysis buffer (25 mM NaF, 1 mM Na3VO4, 50 mM Tris [pH 7.5], 1.5 mM MgCl2, 125 mM NaCl, 0.2% IGEPAL, 5% glycerol, Complete, and RNase Inhibitor). Each clarified lysate was bound overnight to 75 ml of crosslinked HA immunoaffinity resin (Sigma). Unbound proteins were washed off with lysis buffer followed by PBS, and then bound protein complexes were competitively eluted using synthetic HA peptide YPYDVPDYA (250 mg/ml) in PBS. RNA was extracted and a sequencing library prepared using the Illumina Tru-Seq mRNA Library preparation kits.
 
Library strategy OTHER
Library source transcriptomic
Library selection other
Instrument model Illumina HiSeq 2000
 
Description RNA immunoprecipitation (RIP).
Data processing Library strategy: RIP-Seq
Base calling and demultiplexing was performed using CASAVA 1.8.
Alignments were performed using TopHat version 1.4.0 with the following parameters: -p 8 -G ginormous.100806.gtf --no-novel-juncs -a 6 -m 2 --min-intron-length 28 -I 200000 -F 0 -g 1 --library-type fr-unstranded --transcriptome-index ginormous_transcriptome -x 60 -n 2. Ginormous refers to the annotation published in Graveley, B.R., Brooks, A.N., Carlson, J.W., Duff, M.O., Landolin, J.M., Yang, L., Artieri, C.G., van Baren, M.J., Boley, N., Booth, B.W., et al. (2011). The developmental transcriptome of Drosophila melanogaster. Nature 471, 473–479 (PMID 21179090).
bedGraph files were generated from bam files using bedTools with the following parameters: genomeCoverageBed -bg -split
Genome_build: Dm3
Supplementary_files_format_and_content: Genome browser files are presented in bedGraph format.
 
Submission date May 03, 2012
Last update date May 15, 2019
Contact name Brenton R Graveley
E-mail(s) graveley@uchc.edu
Phone 860-679-2090
Organization name University of Connecticut Health Center
Department Genetics and Genome Sciences
Street address 400 Farmington Avenue
City Farmington
State/province CT
ZIP/Postal code 06030-3301
Country USA
 
Platform ID GPL13304
Series (1)
GSE37756 Identification of protein-RNA interactions in Drosophila using RIP-Seq
Relations
SRA SRX145949
BioSample SAMN00990903

Supplementary file Size Download File type/resource
GSM927197_Obar_BRG_Red_FH7121_CG42670_Ps_CAGATC_L007_R1_001_scaled.bedGraph.gz 39.6 Mb (ftp)(http) BEDGRAPH
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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