|
| Status |
Public on Mar 28, 2019 |
| Title |
esggal4NREG80_RNAi_w3_E |
| Sample type |
SRA |
| |
|
| Source name |
intestine
|
| Organism |
Drosophila melanogaster |
| Characteristics |
cell type: intestinal stem cells (ISC)
genotype: esg-gal4ts NREGal80>UAS_RNAi_WHITE
tissue: adult intestine
|
| Growth protocol |
Crosses and adults were kept at 18°C, the GAL80 permissive temperature. 3 day old flies were shifted to 29°C for 2 days to induce RNAi expression
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
RNA was extracted from FACS sorted ISC of 100 females dissected midguts via Arcturus PicoPure RNA isolation Kit and amplified using Arcturus RiboAmp HS plus Kit
RNA libraries were prepared for sequencing using standard Illumina protocols
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Using the damidseq_pipeline reads in fastq files were aligned to the Drosophila melanogaster reference genome version 6 using bowtie2 and alignments were extended to 300 nucleotides or the first GATC site, whichever occurred first.
GATC sites in mappable regions read coverage was counted using bedtools coverage and GATC sites with fewer than 5 counts on average were discarded.
The remaining GATC sites were split into control counts and DamID fusion counts and tested for statistically significant differences using DESeq2, which also estimates a variance stabilized log2 fold enrichment values for each GATC site
Peaks were called by merging 2 or more consecutive significant GATC sites (adjusted p-value < 0.01, log2 fold change > 0). Genes were classified as bound by a protein if 2 consecutive GATC sites within the gene body were occupied with an adjusted p-value < 0.01.
RNA Pol II occupancy was determined by considering mean ratios (Dam-RNA Pol II/Dam-only) across annotated transcripts using “polii.gene.call” script and false discovery rates (FDR) were assigned. Genes with an FDR < 0.01 were used as genes active in ISC.
Reads were quasi-mapped against the Drosophila reference transcriptome fasta using Salmon.
Differential gene expression testing was performed using tximportData, RUVseq.
Genes with an adjusted p-value < 0.01 were considered differentially expressed.
Genome_build: Flybase, release 6.13
|
| |
|
| Submission date |
Mar 27, 2019 |
| Last update date |
Mar 28, 2019 |
| Contact name |
Marius van den Beek |
| E-mail(s) |
m.vandenbeek@gmail.com
|
| Phone |
+33156246580
|
| Organization name |
Institut Curie
|
| Department |
GENETICS AND DEVELOPMENTAL BIOLOGY
|
| Lab |
STEM CELLS AND TISSUE HOMEOSTASIS
|
| Street address |
11-13 rue Pierre et Marie Curie, Equipe Bardin/UMR3215/Batiment BDD
|
| City |
Paris cedex 05 |
| State/province |
Ile-de-France |
| ZIP/Postal code |
75005 |
| Country |
France |
| |
|
| Platform ID |
GPL25244 |
| Series (1) |
| GSE128941 |
Stem cell proliferation is kept in check by the chromatin regulators Kismet/CHD7/CHD8 and Trr/MLL3/4 |
|
| Relations |
| BioSample |
SAMN11266360 |
| SRA |
SRX5582292 |
