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| Status |
Public on Apr 18, 2019 |
| Title |
Retina_p14_ROI |
| Sample type |
SRA |
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| Source name |
eye
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| Organism |
Rattus norvegicus |
| Characteristics |
tissue: retina
condition: hyperoxic (OIR)
strain: Sprague Dawley
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| Extracted molecule |
total RNA |
| Extraction protocol |
TriZol + RNeasy mini Kit (Qiagen, #74106)
Qiaseq miRNA
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| Library strategy |
miRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Basecalls performed using Illumina RTA 2.4.11
Samples demultiplexed using bcl2fastq 2.20
For miRNA entries with identical or near-identical sequences in the miRBase mature database, a manual calibration was created and a new, combined miRNA entry was made for each particular miRNA set. For example, the sequence of hsa-miR-151b is entirely contained within the sequence of hsa-miR-151a-5p. As a result, the miRNA entry is hsa-miR-151b/151a-5p.
Reads are first processed by trimming off the 3’ adapter and low-quality bases using cutadapt (cutadapt.readthedocs.io/en/stable/guide.html). Reads with no adapter sequence are tallied (no_adapter_reads).
Following trimming, the insert sequences and UMI sequences are identified. Reads with less than 16 bp insert sequences (too_short_reads) or less than 10 bp UMI sequences (UMI_defective_reads) are discarded.
To annotate the insert sequences, a unique sequence set is made for all readsets/samples in a submitted job. Following this, a sequential alignment strategy is followed to map to different databases (perfect match to miRBase mature, miRBase hairpin, noncoding RNA, mRNA and otherRNA, and ultimately a second mapping to miRBase mature, where up to two mismatches are tolerated) using bowtie (bowtie-bio.sourceforge.net/index.shtml). At each step, only unmapped sequences pass to the next step. Read counts for each RNA category (miRBase mature, miRBase hairpin, piRNA, tRNA, rRNA, mRNA and otherRNA) are calculated from the mapping results (miRNA_Reads, hairpin_Reads, piRNA_Reads, etc.). miRBase V21 is used for miRNA, and piRNABank is used for piRNA.
A species-specific miRBase mature database is then used, and all remaining unmapped sequences are aligned to the respective genome (RGSC Rnor_6.0) to identify possible novel miRNA molecules.
For each sample, all reads assigned to a particular miRNA or piRNA ID are counted, and the associated UMIs are aggregated to count unique molecules. Read counts and UMI counts are presented in the output Excel ® file “miR_piRNA” sheet. For sequences aligned with tRNAs or otherRNAs, these results are displayed in the “tRNA” or “otherRNA” sheet, respectively. For sequences aligned to genome at the last alignment step, the same information (read counts and clustered UMIs) are output to “notCharacterized_mappable” sheet. Remaining reads are also tallied (notCharacterized_notMappable).
Genome_build: RGSC Rnor_6.0
Supplementary_files_format_and_content: Excel files generated from the pipeline contains a summary tab as well as UMI and read counts for the various miRNAs, pi_RNAs, tRNAs and other small RNAs identified.
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| Submission date |
Apr 17, 2019 |
| Last update date |
Apr 19, 2019 |
| Contact name |
michel desjarlais |
| E-mail(s) |
[email protected]
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| Organization name |
Maisonneuve-Rosemont Hospital Research Center, University of Montréal, Montréal, Québec, Canada.
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| Department |
Department of Ophthalmology,
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| Lab |
Dr sylvain Chemtob
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| Street address |
5415 Blvd L’Assomption
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| City |
montreal |
| State/province |
quebec |
| ZIP/Postal code |
H1T 2M4 |
| Country |
Canada |
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| Platform ID |
GPL20084 |
| Series (1) |
| GSE129995 |
microRNA expression profile in retina and choroid in oxygen-induced retinopathy model |
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| Relations |
| BioSample |
SAMN11457824 |
| SRA |
SRX5706465 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3729063_Retina_p14_ROI.xlsx |
87.2 Kb |
(ftp)(http) |
XLSX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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