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| Status |
Public on Sep 18, 2017 |
| Title |
dsmCherry S2 cells - replicate 4 |
| Sample type |
SRA |
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| Source name |
dsmCherry S2 cells
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| Organism |
Drosophila melanogaster |
| Characteristics |
cell type: S2 cells
cell subtype: dsmCherry S2 cells
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| Growth protocol |
Flies expressing dicer2; asense-Gal4, UAS-CD8::GFP were crossed to flies with empty landing site VIE-260B (VDRC TID:60100) or barc RNAi (VDRC TID:107013) and the progeny was raised at 29C. Late 3rd instar larval brains were dissected and prepared for FACS-sorting. S2 cells were cultured in Schneider's medium + 10% heat-inactivated FBS and treated with dsRNA against mCherry or barc for 7days.
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
Type I neuroblasts were FACS sorted from wild type (TID:60100) or barc RNAi, and RNA was isolated using Trizol LS reagent. S2 cells treated with dsRNA against mCherry or barc were washed and RNA was isolated using Trizol. For both experiments: Total RNA was enriched for polyA mRNA, fragmented and first strand and second strand (dUTP, instead dTTP was used) was generated. For the neuroblast sample, library preparation was done manually using the Illumina Kapa library prep kit, and for the S2 cell sample, library preparation was done with a robot using the Illumina NEB ultra Kit. For both samples, UGDase treatment was done to confer strand specificity.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
Sample S2 2.2 (internal ID 24366)
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| Data processing |
Basecalling RTA 1.18.64.0
The strand specific paired-end reads were screened for ribosomal RNA by aligning with BWA (v0.6.1) against known rRNA sequences (RefSeq). Non matching reads were considerd downstream.
The rRNA subtracted paired-end reads were aligned with TopHat (v1.4.1) against the Drosophila melanogaster genome (FlyBase r5.44) and a maxiumum of 6 missmatches. Introns between 20-150000 bp are allowed which is based on FlyBase statistics. Maximum multihits was set to 1 and InDels as well as Microexon-search was enabled. Additionally, a gene model was provided as GTF (FlyBase r5.44).
Aligned reads in valid pairs are subjected to FPKM estimation with Cufflinks (v1.3.0). In this step bias detection and correction was performed. Furthermore, only those fragments compatible with FlyBase annotation (r5.44) were allowed and counted towards the number of mapped hits used in the FPKM denominator. snRNA, rRNA, tRNA, snoRNA and pseudogenes are masked.
The aligned reads were counted with HTSeq. Counts for technical replicates are summed up. The polyA containing transcripts were subjected to differential expression analysis with DESeq. DESeq parameters were chosen to match the DESeq publication (Anders S, Huber W., 2010).
Genome_build: FlyBase dmel_r5.44
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample (Standard output cufflinks); tab-delimited text file with differentially expressed genes for the whole analysis (standard output DESeq).
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| Submission date |
Mar 13, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Thomas Rainer Burkard |
| E-mail(s) |
thomas.burkard@imba.oeaw.ac.at
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| Organization name |
IMBA
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| Street address |
Dr. Bohrgasse 7
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| City |
Vienna |
| ZIP/Postal code |
1030 |
| Country |
Austria |
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| Platform ID |
GPL17275 |
| Series (1) |
| GSE96549 |
The splicing co-factor Barricade/Tat-SF1, is required for cell cycle and lineage progression in Drosophila neural stem cells. |
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| Relations |
| Reanalyzed by |
GSM3283030 |
| BioSample |
SAMN06564503 |
| SRA |
SRX2636824 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2535391_dsmCherry_24366_3_C6KF5ANXX_genes.fpkm_tracking.txt.gz |
497.8 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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