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Sample GSM2901343 Query DataSets for GSM2901343
Status Public on Apr 01, 2018
Title CLIP_Pcp2_P56_rep1
Sample type SRA
Source name Cerebellar Purkinje cells
Organism Mus musculus
Characteristics cell type: Cerebellar Purkinje cells
age: post-natal day 56
genotype: Pcp2-Cre; Pabpc1-cTag
Extracted molecule total RNA
Extraction protocol cTag-PAPERCLIP on dissected P56 cerebella. Mouse monoclonal anti-GFP clones 19F7 and 19C8 (Heiman et al., 2008) were used for immunoprecipitation.
The PAPERCLIP procedure was performed as previously described (Hwang et al., 2016). Individual cTag-PAPERCLIP libraries were multiplexed and sequenced on MiSeq (Illumina) to obtain 75-nt single-end reads. A barcode sequence was removed from the reads, so the final length of reads is 68-nt.
Library strategy OTHER
Library source transcriptomic
Library selection other
Instrument model Illumina MiSeq
Description PABPC1-bound RNA
Data processing Library strategy: cTag-PAPERCLIP
Analysis of cTag-PAPERCLIP datasets: The processing of raw reads was performed using the CIMS software package as previously described (Moore et al., 2014). Raw reads were filtered based on quality score. Filtered reads with the exact sequence were collapsed into one. Poly(A) sequence at the 3′ end was then trimmed using CutAdapt (Martin, 2011). Only reads that are at least 25-nt in length were mapped to mm10 reference genome. Mapping was performed using Novoalign (Novocraft) without trimming. Reads mapping to the same genomic positions without distinct barcodes were further collapsed into a single read as previously described (Moore et al., 2014). CIMS software package was then used to cluster overlapping collapsed reads from all biological replicates (for each condition) and to determine the number of reads in each cluster. To define a list of high confidence clusters we used the following criteria: clusters had to contain reads from all replicates in each condition, the number of reads in a cluster had to be at least 10% of gene total, clusters had to be located within 20 kb from 3′ ends annotated by RefSeq, clusters that were located upstream of a stretch of 6 or more adenines were excluded due to potential internal priming of the reverse transcription primer. To identify differences in APA between different conditions, Fisher’s exact test was performed comparing the ratio between reads in a particular cluster and the sum of reads in all other clusters in a gene in the two conditions. The output p-values were adjusted for multiple hypotheses testing using the Benjamini-Hochberg method.
Analysis of RNA-seq datasets: RNA-sequencing reads were mapped to the mouse genome (mm10) using Novoalign (Novocraft), reads overlapping genes were counted using HTSeq-count (Anders et al., 2015).
Genome_build: mm10
Supplementary_files_format_and_content: Columns in tab-delimited files for CLIP datasets represent the following: name_cluster (name of a CLIP cluster), id_gene (Refseq gene ID), chr_cluster (chromosome where cluster is located), start_cluster (cluster start), end_cluster (cluster end), strand_cluster (strand), name2 (gene symbol) and number of reads from each replicate in a given cluster.
Supplementary_files_format_and_content: Tab-delimited text files for RNA-seq datasets represent Refseq gene ID and number of reads overlapping with exons.
Submission date Dec 22, 2017
Last update date Apr 01, 2018
Contact name Saša Jereb
Organization name The Rockefeller University
Department Laboratory of Molecular Neuro-Oncology
Lab Robert B. Darnell Lab
Street address 1230 York Ave
City New York
State/province New York
ZIP/Postal code 10065
Country USA
Platform ID GPL16417
Series (1)
GSE108480 Differential 3’ Processing of Specific Transcripts Expands Regulatory and Protein Diversity Across Neuronal Cell Types
BioSample SAMN08235545
SRA SRX3506795

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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