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Sample GSM7059844 Query DataSets for GSM7059844
Status Public on Sep 26, 2023
Title Dana.ovary_FC.1 small-RNA-Seq
Sample type SRA
 
Source name Dana.ovary_FC.1
Organism Drosophila ananassae
Characteristics tissue: ovary
enrichment: somatic follicle cells
assembly: d101g,d15genomes,droAna2,GCF_003285975,GCF_017639315
Treatment protocol N/A
Growth protocol All Drosophila species were maintained at room temperature on species-specific food.
Extracted molecule total RNA
Extraction protocol Small RNAs were isolated using the TraPR Small RNA Isolation Kit (Lexogen) following the manufacturer's instructions. The soma-enrichment sRNA-seq libraries were generated for 13 species (2 replicates each) similar to published protocols (Akkouche et al., 2013; Chirn et al., 2015), with modifications. In brief, 75-100 ovary pairs were dissected in ice-cold PBS. Ovaries were dissociated for 18 minutes in 0.25% Trypsin (Sigma-Aldrich) at 25˚C, shaking at 800 rpm. Dissociated tissue was pushed through a 40 µm nylon mesh (Greiner Bio-one) washed with equal volume Schneider 2 medium (Thermo Fisher Scientific) and then pelleted. Cell count was quantified using the Luna-FL (Logos Biosystems). Cells were pelleted and directly used as input for sRNA isolation using the TraPR Small RNA Isolation Kit (Lexogen).
sRNA libraries were generated using the Small RNA-Seq Library Prep Kit (Lexogen) with minor modifications. Both primers A3 and A5 as well as the primer RTP were used at 0.5x.
 
Library strategy ncRNA-Seq
Library source transcriptomic
Library selection size fractionation
Instrument model Illumina NovaSeq 6000
 
Description Dana.ovary_FC.unox.SLX-22089-1.1
Data processing Trim Galore! (v0.6.4, --stringency 30 -e 0.1 -a TGCTTGGACTACATATGGTTGAGGGTTGTA --length 18 -q 0) was first run to remove an abundant rRNA sequence, followed by a second run (--stringency 5 -e 0.1 --length 18 --max_length 35 -q 0) to remove adapter sequences (specified using ‘-a’), and any flanking random nucleotides (‘--clip_R1’ and/or ‘--three_prime_clip_R1’ with appropriate arguments).
The processed reads were mapped to a miRNA hairpin database (miRBase release 22.1) using bowtie (v1.2.3, -S -n 2 -M 1 -p 20 --best --strata --nomaqround --chunkmbs 1024) with ‘--un’ and ‘--max’ to extract unmapped reads.
Reads not mapping to miRNAs were aligned to the respective reference genomes using bowtie (-S -n 2 -M 1 -p 20 --best --strata --nomaqround --chunkmbs 1024).
Multi-mapping reads were extracted into a separate BAM file using awk (MQ<10).
The BAM files were converted to bigWig using bamCoverage from deepTools (Ramírez et al., 2016) (v3.3.2, --binSize 1 --ignoreForNormalization chrM --normalizeUsing CPM --exactScaling --skipNonCoveredRegions --minFragmentLength 23 --maxFragmentLength 30) and additionally ‘--filterRNAstrand’ to separate the two strands, ‘---scaleFactor’ to scale counts per million to reflect all mapped reads, and optionally ‘--minMappingQuality 50’ when extracting uniquely mapped reads.
Supplementary files format and content: bigWig showing read coverage per position. File name indicate species, genome assembly, strand, and sample.
 
Submission date Feb 22, 2023
Last update date Sep 26, 2023
Contact name Susanne Bornelöv
E-mail(s) susanne.bornelov@cruk.cam.ac.uk
Organization name University of Cambridge
Department Cancer Research UK - Cambridge Institute
Lab Hannon
Street address Robinson Way
City Cambridge
ZIP/Postal code CB2 0RE
Country United Kingdom
 
Platform ID GPL33163
Series (2)
GSE225888 Unistrand piRNA clusters are an evolutionarily conserved mechanism to suppress endogenous retroviruses across the Drosophila genus [small RNA-Seq]
GSE225889 Unistrand piRNA clusters are an evolutionarily conserved mechanism to suppress endogenous retroviruses across the Drosophila genus
Relations
BioSample SAMN33419777
SRA SRX19478060

Supplementary file Size Download File type/resource
GSM7059844_Dana.GCF_003285975.minus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 2.1 Mb (ftp)(http) BW
GSM7059844_Dana.GCF_003285975.plus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 1.7 Mb (ftp)(http) BW
GSM7059844_Dana.GCF_017639315.minus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 2.0 Mb (ftp)(http) BW
GSM7059844_Dana.GCF_017639315.plus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 1.7 Mb (ftp)(http) BW
GSM7059844_Dana.d101g.minus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 2.2 Mb (ftp)(http) BW
GSM7059844_Dana.d101g.plus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 1.8 Mb (ftp)(http) BW
GSM7059844_Dana.d15genomes.minus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 2.1 Mb (ftp)(http) BW
GSM7059844_Dana.d15genomes.plus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 1.8 Mb (ftp)(http) BW
GSM7059844_Dana.droAna2.minus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 2.5 Mb (ftp)(http) BW
GSM7059844_Dana.droAna2.plus.ovary_FC.unox.SLX-22089-1.uniq.23-30.bw 2.2 Mb (ftp)(http) BW
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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