|
| Status |
Public on Oct 28, 2020 |
| Title |
Young adults purified DNA muscle-specific DamID GFP control, replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
Purified DNA
|
| Organism |
Caenorhabditis elegans |
| Characteristics |
developmental stage: Young adults
tissue: Muscle
sample type: Specific expression of Dam in target tissue
strain: PMW748
expressing: Dam GFP in muscle cells
|
| Growth protocol |
Worms were grown on NGM seeded with Dam negative E. coli GM48 for at least two generations. Around 5,000 synchronized L1s were seeded onto 100 mm plates (1,000-1,200 per plate) and collected 53h later. Worms were washed extensively with M9 (at least 10 times) and distributed in aliquots of 30 μl removing the excess of liquid. Samples were snap-frozen and stored at −80 °C.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
gDNA was extracted from the animals using DNeasy Blood and Tissue Kit (Qiagen #69504).
DamID amplicons were obtained as previously described (Gomez-Saldivar et al. 2016) with 20 PCR cycles for worm-wide DamID, 22 PCR cycles for muscle DamID, 22-24 for intestine DamID and 24-26 cycles for XXX cells DamID.
DamID PCR amplicons were directly used for nanopore library barcoding and preparation using NBD-104 and LSK-109 kits (Oxford Nanopore).
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
MinION |
| |
|
| Description |
DamID amplicons from total DNA of animals expressing Dam GFP in muscle cells
Muscle_DamID_pass_barcode12.sorted_damid-vs-Dam.gatc.bedgraph
Muscle_DamID_pass_barcode12.genes.details.csv
|
| Data processing |
Library strategy: DamID
Nanopore sequences were basecalled and demultiplexed using guppy 3.6 with the high accuracy model before mapping to the ce11 genome using minimap2 (Li 2018).
Reads were considered as DamID amplicons when both ends mapped +/- 8 bp from a genomic GATC motif. The bam files represent the filtered reads.
Filtered libraries were then used to call polymerase footprinting values using the damidseq_pipeline package (O. J. Marshall and Brand 2015) using bam files (parameters: --bamfiles --extend_reads=0). The output of this step are bedgraphs.
Gene values were extracted using polii.gene.call (https://owenjm.github.io/damidseq_pipeline/) using WS270 genes definition. The output of this steps are csv files.
Genome_build: ce11
Supplementary_files_format_and_content: bam file with valid DamID reads
Supplementary_files_format_and_content: Normalized begraph (Dam fusion over Dam only)
Supplementary_files_format_and_content: Gene-level DamID values in csv format, with number of GATC per gene and FDR
|
| |
|
| Submission date |
Sep 03, 2020 |
| Last update date |
Oct 29, 2020 |
| Contact name |
Peter Meister |
| E-mail(s) |
peter.meister@unibe.ch
|
| Phone |
+41316844609
|
| Organization name |
University of Bern
|
| Department |
Institute of Cell Biology
|
| Lab |
Cell Fate and Nuclear Organization
|
| Street address |
Baltzerstrasse 4
|
| City |
Bern |
| State/province |
Schweiz |
| ZIP/Postal code |
3012 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL26522 |
| Series (2) |
| GSE157416 |
Tissue-specific transcription footprinting using RNA PolII DamID (RAPID) in C. elegans [Young adult] |
| GSE157418 |
Tissue-specific transcription footprinting using RNA PolII DamID (RAPID) in C. elegans |
|
| Relations |
| BioSample |
SAMN16049234 |
| SRA |
SRX9067989 |
