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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Sep 13, 2017 |
| Title |
cTag-PAPERCLIP for Cx3cr1-Cre; Ctag-PABP mouse brain, replicate 1 |
| Sample type |
SRA |
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| Source name |
mouse brain
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| Organism |
Mus musculus |
| Characteristics |
sample type: replicate 1
tissue: brain
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| Extracted molecule |
total RNA |
| Extraction protocol |
Sample preparation, immunoprecipitation, SDS-PAGE and RNA extraction were performed using the PAPERCLIP protocol (Hwang et al. 2016). Mouse monoclonal anti-GFP antibodies clone 19F7 and 19C8 (Heiman et al. 2014) were used for immunoprecipitation.
The sequencing library was constructed using the PAPERCLIP method (Hwang et al. 2016). The library contains a 11-nt degenerate linker sequence at the 5′ end (3-nt degenerate sequence, 4-nt sample multiplexing index and 7-nt random barcode).
Other (cTag-PAPERCLIP)
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
Index 3: CGAT
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| Data processing |
The processing of raw reads was performed using the CIMS software package previously described (Moore et al. 2014). In brief, the raw reads were filtered based on quality score.
Filtered reads with the exact sequence were collapsed into one.
The 5′ degenerate linker was removed.
Poly(A) sequence at the 3′ end was then trimmed using CutAdapt (Martin 2011). Only reads that are at least 25-nt in length are retained for mapping to reference genome (mm10).
Mapping was performed using Novoalign (http://www.novocraft.com) without trimming. A minimum of 25-nt match to the genome sequence was required and only those reads mapped unambiguously to the genome (single hits) were kept for downstream analysis.
Reads mapping to the same genomic positions without distinct barcodes were further collapsed into a single tag as previously described (Moore et al. 2014). All reads went through the entire process are referred to as “unique reads” and were used for poly(A) site annotation and other downstream analysis.
Unique reads from both replicates were merged. CIMS software package was used to cluster overlapping reads and to determine the read counts (PH) for each cluster. The clusters were filtered by PH before downstream analysis. For each filtered cluster, the most abundant 3′ end from all reads in the cluster was assigned as the poly(A) site. The 3′ most position was selected if there was a tie in abundance for multiple 3′ ends. Please refer to the citation for further information.
Note: The submitted raw files are de-multiplexed and quality-filtered without further processing.
Genome_build: mm10
Supplementary_files_format_and_content: txt files for unique read counts at gene level (RefSeq)
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| Submission date |
Jan 25, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Hun-Way Hwang |
| E-mail(s) |
Hunway.Hwang@pitt.edu
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| Organization name |
University of Pittsburgh
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| Department |
Department of Pathology
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| Lab |
S754 Scaife Hall
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| Street address |
3550 Terrace Street
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| City |
Pittsburgh |
| State/province |
PA |
| ZIP/Postal code |
15261 |
| Country |
USA |
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| Platform ID |
GPL17021 |
| Series (1) |
| GSE94054 |
cTag-PAPERCLIP Reveals Alternative Polyadenylation Promotes Cell-Type Specific Protein Diversity and Shifts Araf Isoforms with Microglia Activation |
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| Relations |
| BioSample |
SAMN06269366 |
| SRA |
SRX2515129 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2467574_mm10.hh.gene.meta.RefSeq.V2.sorted.tobed6.intersect.B5p10.No3.count.sort.txt.gz |
74.6 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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