|
| Status |
Public on Jul 25, 2023 |
| Title |
imb_keller_2022_03_12_IgG_empty_Rep2 |
| Sample type |
SRA |
| |
|
| Source name |
Ag55 cell line
|
| Organism |
Anopheles gambiae |
| Characteristics |
cell line: Ag55 cell line
antibody: IgG control Rabbit (Abcam ab37415)
ectopic expression: HA-tagged empty control
|
| Treatment protocol |
2 Mio. Ag55 cells were seeded in a 6-well plate. After 16 hours, 600 μl of a selected baculovirus in Sf-900 media was added to the cells. After 6 hours the media was changed to fresh L15. Cells were collected for further processing 48h from the addition of the baculovirus.
|
| Growth protocol |
Ag55 cells were cultured in Leibovitz L15 media with 10% FBS (Gibco, 10270-10,6 Lot: 2260092) and 1x penicillin-streptomycin (Gibco, 15140122) at 27°C at 80% humidity.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The cell suspension was counted and 0.4 Mio cells were used for each reaction. The experiment was performed following the native protocol dx.doi.org/10.17504/protocols.io.bcuhiwt6. Antibodies are listed with the respective samples. We used pA-Tn5 prepared by the IMB protein production core facilty.
CUT&Tag libraries were amplified using NEBNext Ultra II Q5 Master Mix with standard protocols and 15 PCR cycles. Libraries were cleaned up using 1.3 volume Ampure XP beads and pooled together for paired-end sequencing.
OTHER: CUT&Tag
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
IgG_empty_Rep2
|
| Data processing |
Reads were trimmed with cutadapt (v1.18) to remove Illumina adapter sequences.
Trimmed reads were mapped to the ensembl AgamP4 genome with bowtie2 (v2.3.4)
Coverage signal tracks (bigWigs) of alignments were generated using Samtools (v1.9) and DeepTools (v3.1)
Peaks were called with MACS2 (v2.1.2) using the corresponding IgG samples as controls.
A greylist was generated from IgG samples and applied for peak filtering
DiffBind (v3.0.5) was use to determine differential binding, with normalization to background bins (15000 bp). Only peaks with FDR<0.05 in were considered as differentially bound.
Assembly: AgamP4
Supplementary files format and content: bigWig files, containing CPM normalized coverage signals of primary alignments
|
| |
|
| Submission date |
Aug 05, 2022 |
| Last update date |
Jul 25, 2023 |
| Contact name |
Frank Rühle |
| E-mail(s) |
f.ruehle@imb-mainz.de
|
| Organization name |
Institute of Molecular Biology (IMB)
|
| Department |
Bioinformatics Core Facility
|
| Street address |
Ackermannweg 4
|
| City |
Mainz |
| ZIP/Postal code |
55128 |
| Country |
Germany |
| |
|
| Platform ID |
GPL22033 |
| Series (2) |
| GSE210627 |
SOA confers X chromosome dosage compensation in Anopheles gambiae mosquitoes (CUT&Tag I) |
| GSE210630 |
SOA confers X chromosome dosage compensation in Anopheles gambiae mosquitoes |
|
| Relations |
| BioSample |
SAMN30167116 |
| SRA |
SRX16881232 |
