|
| Status |
Public on May 24, 2024 |
| Title |
SuHw_LOF_ovaries_1_8st_ChIP_Rpb3 |
| Sample type |
SRA |
| |
|
| Source name |
Ovaries were dissected from females 15 hours after eclosion (containing egg chambers corresponding to 1-8 stages)
|
| Organism |
Drosophila melanogaster |
| Characteristics |
tissue: Ovaries were dissected from females 15 hours after eclosion (containing egg chambers corresponding to 1-8 stages)
chip antibody: anti-Rpb3 antibodies (Mazina et al (2018) Biochim. Biophys. Acta BBA - Gene Regul. Mech. 1861, 178–189)
genotype: su(Hw)v/E8 stock (a gift of A. Golovnin)
treatment: no treatment
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
For the ChIP-Seq experiments ovaries were collected in PBS buffer (50 pairs per ChIP), cross-linked with 1% of formaldehyde for 5 min and then incubated for 5 min with 125 mM Glycine. After that ovaries were washed with PBS buffer for three times. The remaining ChIP protocol was performed as described previously (Vorobyeva et al (2013) NAR, 41:11, 5717–5730). For the in situ Hi-C experiments, ovaries were collected in PBS buffer (200 pairs per Hi-C), cross-linked with 1% of formaldehyde for 5 min and then incubated for 5 min with 125 mM Glycine. Then ovaries were washed with 1xPBS buffer for three times. The remaining in situ Hi-C protocol was performed as described previously (Rao et al (2014) Cell, 159:1665-1680). For extraction of RNA the ovaries of 15 hour old wild-type (oregon) and su(Hw)v/E8 females were collected in PBS buffer (50 pairs per sample), in two biological repeats. Total RNA was extracted with the TRI reagent (Ambion). PolyA comprising RNA fraction was isolated and prepared for sequencing with the NEBNext UltraTM II Directional RNA Library Prep Kit.
ChIP-Seq, Hi-C and RNA-Seq libraries were prepared for sequencing using standard Illumina protocols
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
bcl2fastq v2.17.1.14 Conversion Software (Illumina)
For ChIP-Seq data the single‐end reads in FastQ format were mapped to the Drosophila genome assembly dm6 using Bowtie2 (Galaxy Version 2.3.4.3) and filtered (with minimum MAPQ quality score = 5). BigWig files were generated using bamCoverage (Galaxy Version 3.5.1.0.0) with scores representing number of reads per kilobase per million (RPKM). The final BigWig files (representing the protein-binding profiles) were obtained using bigwigcompare (Galaxy Version 3.5.1.0.0) as a ratio of ChIP signal to Input.
For RNA-Seq data the paired-end reads were mapped to the Drosophila genome assembly dm6 using HISAT2 (Galaxy Version 2.2.1+galaxy0). BigWig files were generated using bamCoverage (Galaxy Version 3.3.2.0.0) with scores representing number of reads per kilobase per million (RPKM).
Hi-C data was processed using the HiCExplorer suite version 3.4.2. The paired‐end reads in FastQ format were separately mapped to the Drosophila genome assembly dm6 using Bowtie2 (Galaxy Version 2.4.2+galaxy0), with the following options: --sensitive-local --reorder. The raw contact Hi-C matrices in .h5 format with the bins size of 4 kb were generated in hicBuildMatrix (Galaxy Version 3.4.2.0).
Assembly: dm6 (Drosophila melanogaster Release 6 plus ISO1 MT)
Supplementary files format and content: bigwig
|
| |
|
| Submission date |
May 03, 2023 |
| Last update date |
May 24, 2024 |
| Contact name |
Marina Yu. Mazina |
| E-mail(s) |
mazinam@genebiology.ru
|
| Phone |
+79771075035
|
| Organization name |
Institute of Gene Biology RAS
|
| Street address |
Vavilova, 34/5
|
| City |
Moscow |
| ZIP/Postal code |
119334 |
| Country |
Russia |
| |
|
| Platform ID |
GPL25244 |
| Series (1) |
| GSE231576 |
Su(Hw) interacts with Combgap to establish long-range chromatin contacts |
|
| Relations |
| BioSample |
SAMN34569546 |
| SRA |
SRX20209940 |
