|
| Status |
Public on Aug 10, 2022 |
| Title |
Dam2 |
| Sample type |
SRA |
| |
|
| Source name |
medulla neuroblasts from larval brain
|
| Organism |
Drosophila melanogaster |
| Characteristics |
cell type: Drosophila medulla neuroblasts
treatment: Induce at 29 C for 72 hours
genotype: UAS-Dcr2; tubG80ts/CyO; SoxN-Gal4/UAS-Dam
|
| Treatment protocol |
The expressions of the Dam-fusion proteins were then induced in larval brains by shifting the larvae to 29°C for 72 hours. Climbing third instar larvae were then dissected. Only the optic lobes were used for further sample preparation. For each sample around 100 brains were dissected.
|
| Growth protocol |
Mating crosses were incubated at 25 °C. Eggs laid within an 8-hour window were then shifted to 18°C for 3 days until they hatched.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
fly brains were dissected in 1X PBS and genomic DNA was isolated using QIAamp DNA Micro kit (Qiagen). Genomic DNA was then digested with DpnI and purified using spin column purification (Qiagen PCR-purification kit). To the DpnI digested DNA, DamID-PCR adaptors were ligated. Subsequently the adaptor ligated DNA fragments were digested with DpnII. The DpnII digested genomic DNA fragments were then amplified by PCR to enrich for bound GATC fragments.
To make sequencing libraries the PCR enriched genomic DNA sample was sonicated using a Bioruptor bath sonicator (Diagenode), purified using AMPure XP beads, end-repaired and 3’-adenylated. Illumina sequencing indexes were then ligated to these fragments. Index-labeled DNA fragments were then further enriched by PCR amplification, checked for quality and fragment size distribution on an Agilent Bioanalyzer and by qPCR.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Library strategy: Dam-ID-seq
DamID-seq data was analyzed using the damidseq-pipeline(Marshall, O. J. & Brand, A. H. Bioinformatics 31, 3371-3373, doi:10.1093/bioinformatics/btv386 (2015)). Duplicate samples of Ey-Dam and Su(H)-Dam samples and Dam-only controls were aligned to the Drosophila reference genome UCSC dm6.
The main utility of the damid-seq pipeline was used to align reads to the genome using bowtie2, bin and count reads, normalize counts and generate log-2 ratio bedgraph files for each DamID-sample and its corresponding Dam-only control. . The provided gatc.track file was used for running the script.
Next, the findpeaks utility was used with an F.D.R. <0.01 to identify peaks.
We then used the provided peaks2genes utility to assign peaks to genes.
To assess the reproducibility of our data we also ran the findpeaks script using F.D.R <0.1 to discover peaks with weaker statistical confidence and used this as input for the I.D.R. Python package (https://github.com/nboley/idr). 984 of 1810 (54.4.%) Ey-Dam and 972 of 1996 (48.7%) of Su(H)-Dam peaks passed an I.D.R. threshold of 0.05. Lists of genes bound by both replicates of Ey-Dam and by Su(H)-Dam were used in BioVenn89 to visualize the overlap between these two lists and estimate overlap percentage.
Genome_build: UCSC dm6
Supplementary_files_format_and_content: Ey_1-vs-Dam.gatc-FDR0.01.peaks.gff: gff file showing locations of peaks with FDR<0.01 for EyDam1
Supplementary_files_format_and_content: Ey_2-vs-Dam.gatc-FDR0.01.peaks.gff: gff file showing locations of peaks with FDR<0.01 for EyDam2
Supplementary_files_format_and_content: Ey_overlap_all_replicate_peaks.gff: gff file showing overlaps of FDR<0.01 peaks for two EyDam replicates
Supplementary_files_format_and_content: Ey_and_SuH_overlap_allpeaks.gff: gff file showing overlaps of Ey binding peaks and Su(H) bidnings peaks
Supplementary_files_format_and_content: SuH_1-vs-Dam.gatc-FDR0.01.peaks.gff: gff file showing locations of peaks with FDR<0.01 for SuHDam1
Supplementary_files_format_and_content: SuH_2-vs-Dam.gatc-FDR0.01.peaks.gff: gff file showing locations of peaks with FDR<0.01 for SuHDam2
Supplementary_files_format_and_content: SuH_overlap_all_replicate_peaks.gff: gff file showing overlaps of FDR<0.01 peaks for two SuHDam replicates
|
| |
|
| Submission date |
Nov 11, 2021 |
| Last update date |
Aug 10, 2022 |
| Contact name |
Xin Li |
| E-mail(s) |
lixin@illinois.edu
|
| Phone |
2172443784
|
| Organization name |
University of Illinois Urbana-Champaign
|
| Department |
CDB
|
| Lab |
Li
|
| Street address |
601 S. Goodwin Avenue, B107 CLSL
|
| City |
Urbana |
| State/province |
IL |
| ZIP/Postal code |
61801 |
| Country |
USA |
| |
|
| Platform ID |
GPL25244 |
| Series (1) |
| GSE188643 |
A Notch-dependent transcriptional mechanism controls expression of temporal patterning factors in Drosophila medulla |
|
| Relations |
| BioSample |
SAMN23075006 |
| SRA |
SRX13114377 |
