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Sample GSM1091908 Query DataSets for GSM1091908
Status Public on Aug 19, 2013
Title Input-PITX1
Sample type SRA
 
Source name sonified chromatin from Pitx1(wt) expressing chMM
Organism Gallus gallus
Characteristics cell type: chicken micromass
days of incubation: 9
virus used for infection: RCASBP(A)-3xFLAG-cPITX1(wt)
chip antibody: none
Growth protocol Chicken micromass cultures were routinely prepared by generating a single cell suspension of embryonic mesenchymal chicken limb bud cells (HH24) (2 × 10^7 cells/ml) and the appropriate virus was added before plating. Cultures were plated as 10µl droplets and cells were cultured for 9 days in chMM medium (DMEM/HAM’S F12 (Gibco) with 10% FBS (PAA), 0.2% chicken serum (Sigma), 1% L-glutamin (Gibco), 1% penicillin/streptomycin (Gibco) before harvest.
Extracted molecule genomic DNA
Extraction protocol Lysates were clarified from sonicated nuclei and protein-DNA complexes were enriched with antibody. For ChIP-seq library preparation ethanol precipitated DNA from ChIP was redissolved in 46 µl of water and subjected to library preparation using the NEBnext ChIP-Seq Library Prep Master Mix for Illumina (New England Biolabs) selecting for a fragment size range of 300-450 bp according to manufacturer's instructions. Sequencing was performed on an Illumina GAIIx sequencer with 36 bp single reads using TruSeq v5 sequencing chemistry and TruSeq v5 Cluster Generation Kits. For RNA-seq, total RNA from chicken micromass cultures was isolated using peqGOLD TriFast Reagent (peqLAB) and subsequent purification with RNeasy Mini Kit (Qiagen) according to manufacturer's instructions. mRNA for RNA-seq libraries was purified using the Oligotex mRNA Mini Kit (Qiagen) from 6 µg of total RNA. RNA-seq libraries were constructed using the NEBNext mRNA Library Prep Master Mix Set for Illumina according to manufacturer's instructions selecting for a fragment size range of 300-500 bp. Sequencing was performed on an Illumina GAIIx sequencer with 115 bp single reads using TruSeq v5 sequencing chemistry and TruSeq v5 Cluster Generation Kits.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina Genome Analyzer IIx
 
Data processing Peaks were called using MACS2 with appropriate input controls and the following settings: ( -g 1. 0e9 -q None -p 1e-1 --too-large --bw (as determined by spp-get binding characteristics))
 
Submission date Mar 01, 2013
Last update date May 15, 2019
Contact name Peter Hansen
E-mail(s) peter.hansen@charite.de
Phone +49 (0)30 450-569127
Organization name Charité-Universitätsmedizin Berlin
Department Institute for Medical Genetics and Human Genetics
Lab Computational Biology Group
Street address Augustenburger Platz 1
City Berlin
ZIP/Postal code 13353 Berlin
Country Germany
 
Platform ID GPL13797
Series (1)
GSE44799 Distinct Global Shifts in Genomic Binding Profiles of Limb Malformation Associated HOXD13 Mutations
Relations
SRA SRX248428
BioSample SAMN01974726

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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