|
| Status |
Public on Dec 27, 2022 |
| Title |
HisC, 6.5-7h, STRIPEseq, rep5 |
| Sample type |
SRA |
| |
|
| Source name |
whole embryo
|
| Organism |
Drosophila melanogaster |
| Characteristics |
tissue: whole embryo
cell type: Drosophila embryo 6.5-7h AEL
genotype: Df(2L)HisC, P{GAL4-twi.2xPE}/ Df(2L)HisC, P{UAS:eYFP}AH2
|
| Treatment protocol |
Maintaining flies used for embryo collection: controlled light conditions (24h light on), new apple agar plates were provided with yeast twice a day, raised at 25°C
Embryos were washed with PBST and dechorionated in 50% bleach, mutant embryos were identified by eYFP expression
|
| Growth protocol |
Experimental design: Flies were raised on apple agar petri dishes in acrylic cages for 30 minutes for egg laying at 25°C, petri dishes were exchanged with new ones and embryos were further incubated at 25°C for 5.5, 6.5 hours
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total mRNA was extracted using fresh or with RNAlater (Invitrogen) preserved (at -80°C) embryos. mRNA is extracted using Qiagen RNeasy Plus Micro kit adjusting the protocol for small RNA species as described by the manufacturer
200ng total input RNA was used. Uncapped mRNA was digested by Terminator Exonuclease. Adapter and indices were added by template switching oligo PCR, the final library was amplified by PCR using NEB Ultra II Q5 PCR master mix M0544S (14 cycles) following the protocol published by Policastro et al., 2020
STRIPE-seq, Policastro et al., 2020
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
5'capped mRNA
|
| Data processing |
Quality of STRIPEseq data sets was analyzed using FastQC v0.11.5
STRIPEseq read files were processed and aligned to Drosophila melanogaster reference genome dm6 (FlyBase Dmel Release 6.23) following the GoSTRIPES workflow (https://github.com/BrendelGroup/GoSTRIPES)
Reads counts were further calculated and TSSs were called using TSRchitect (https://www.bioconductor.org/packages/release/bioc/html/TSRchitect.html). The threshold for a TSS to be called was set to at least 5 raw counts that had to cluster into a TSR consistently in at least three of the analyzed replicates
Read counts were then normalized and differential TSR analysis was performed using DESeq2 with default settings. TSR shape analysis was accomplished using TSRexplorer. Annotation of TSSs and TSRs was done with ChIPseeker using a promoter window from -/+ 250bp of a TSS
Genome browser tracks in bigwig format were generated from merged replicates using deeptools bamCoverage
Assembly: dm6
|
| |
|
| Submission date |
May 20, 2022 |
| Last update date |
Dec 27, 2022 |
| Contact name |
Ufuk Günesdogan |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Göttingen
|
| Street address |
Justus-von-Liebig Weg 11
|
| City |
Göttingen |
| ZIP/Postal code |
37077 |
| Country |
Germany |
| |
|
| Platform ID |
GPL25244 |
| Series (2) |
| GSE203477 |
Recycling of parental histones preserves the epigenetic landscape during embryonic development (STRIPE-seq) |
| GSE203478 |
Recycling of parental histones preserves the epigenetic landscape during embryonic development |
|
| Relations |
| BioSample |
SAMN28569321 |
| SRA |
SRX15392705 |
