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Sample GSM1269955 Query DataSets for GSM1269955
Status Public on Nov 21, 2013
Title n2a-lacZ-R1
Sample type SRA
Source name N2A_lacZ
Organism Mus musculus
Characteristics cell line: neuro 2A (N2A)
cell type: neuroblastoma
molecule subtype: genomic DNA associated with a control oligonucleotide (corresponding to the E.coli LacZ sequence)
Treatment protocol None.
Extracted molecule genomic DNA
Extraction protocol CHART Enrichment and RNase H Mapping experiments were performed as previously described (Simon, 2013, Capture hybridization analysis of RNA targets (CHART). Curr Protoc Mol Biol Chapter 21: Unit 21 25). CHART extract was prepared from approximately 8 x 107 N2A cells per pull down and hybridized with 810 pmol biotinylated oligonucleotide cocktail overnight with rotation at room temperature. Complexes were captured using 250ml MyOneC1 streptavidin beads (Invitrogen) overnight at room temperature with rotation. Bound material was extensively washed and eluted using RNase H (New England Biolabs) for 30 min at room temperature. Samples were treated with Proteinase K and cross-links were reversed. DNA was sheared to an average fragment size of 150-300 bp using a Bioruptor (Diagenode).
Fragmented DNA was end-repaired, A-tailed and adapter-ligated before size selection and amplification. The prepared libraries were QC’ed and multiplexed before paired end sequencing over one lane of a Illumina HiSeq flow cell. For more information contact the High-Throughput Genomics Group at the Wellcome Trust Centre for Human Genetics, Oxford.
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina HiSeq 2500
Description LacZ1
Data processing library strategy: Capture Hybridisation Analysis of RNA Targets (CHART)-seq
alignment: 50bp, paired-end reads were mapped to the mouse genome (mm9) using bowtie with the options “-m1 –v2 –best –strata –a”
peak-calling: Peak calls were made using the MACS2 algorithm (Zhang Y, Liu T, Meyer CA, Eeckhoute J, Johnson DS, Bernstein BE, Nusbaum C, Myers RM, Brown M, Li W, Liu XS (2008) Model-based analysis of ChIP-Seq (MACS). Genome Biol 9: R137) ( with the options “--mfold 10 30 --gsize=2.39e9 --qvalue=0.01”. Peak calls were then filtered such that only peak calls with a -log10 qvalue > 5 were retained (FDR < 0.001%). Peaks were called seperately for (i) Paupar1 (treatment) vs LacZ1 (control), (ii) Paupar2 (treatment) vs LacZ2 (control), (iii) Paupar1 (treatment) vs input1 (control), (iv) Paupar2 (treatment) vs Input1 (control).
construction of final peak set: The final peakset comprises of peaks common to all four individual comparisions. Overlapping peaks were merged in this analysis.
Genome_build: mm9
Supplementary_files_format_and_content: A single bed file containing the final paupar peak set. The average MACS2 peak score is reported in the score column.
Submission date Nov 20, 2013
Last update date May 15, 2019
Contact name Stephen Sansom
Organization name Kennedy Institute of Rheumatology
Department NDORMS
Lab Sansom
Street address Roosevelt Drive
City Oxford
State/province Oxfordshire
ZIP/Postal code OX3 7FY
Country United Kingdom
Platform ID GPL17021
Series (2)
GSE52570 The long non-coding RNA Paupar regulates the expression of both local and distal genes [CHART-seq]
GSE52571 The long non-coding RNA Paupar regulates the expression of both local and distal genes
BioSample SAMN02415144
SRA SRX380397

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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