|
| Status |
Public on Oct 09, 2019 |
| Title |
neural crest bulk TF_KD rep2 RNA-seq |
| Sample type |
SRA |
| |
|
| Source name |
dorsal neural tube microdissection
|
| Organism |
Gallus gallus |
| Characteristics |
developmental stage: stage 5-10ss
tissue: neural crest
isolation_method: embryo microdissection
amplification: SMART-seqTMv4
genotype: TF knockdown
|
| Treatment protocol |
Dissected cranial regions from electroporated embryos were dissociated with dispase (1.5mg/ml in DMEM/10mM Hepes pH 7.5) at 37°C for 15min with intermittent pipetting to achieve a single cell suspension and 0.05% Trypsin at 37°C for a final 3min. The reaction was stopped and cells were resuspended in an excess of Hanks buffer. Cells were centrifuged at 500g for 10min and resuspended in Hanks buffer, passed through a 0.22mm filter and centrifuged at 750g for 10min, pelleted cells were resuspended in 500µl Hanks buffer. Fluorescent positive cells were sorted and collected using BD FACS-Aria Fusion. We collected ~300 and ~600 NC cells per embryo at 5-6ss and 8-10ss respectively.
|
| Growth protocol |
Fertilised wild-type chicken eggs were obtained from Henry Stewart & Co (Norfolk), staged according to Hamburger and Hamilton (1951) (24). All experiments were performed on chicken embryos younger than 12 days of development, and as such were not regulated by the Animals (Scientific Procedures) Act 1986.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted from dissected half dorsal neural tubes using Ambion RNAqueous Micro Total RNA isolation kit (Cat.#AM1931, ThermoFisher Scientific), the integrity was checked using Bioanalyser, and only samples with RIN>7 were used for analysis. 6 individual embryos and associated RNA-seq libraries were analysed.
RNA-seq libraries were prepared using SMART-Seq™ v4 Ultra™ Low Input RNA Kit for Sequencing (Cat.#634889, Takara Bio Clontech) and sequenced using 40bp paired-end reads on Illumina NextSeq500.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
TFs_RNA-seq.txt
|
| Data processing |
Reads were mapped using STAR (v.2.4.2a) for RNA-seq to galGal5. Read counts were obtained using subread (v.1.4.5) FeatureCounts
Differential Expression analysis was performed using DESeq2 R package.
Genome_build: galGal5
Supplementary_files_format_and_content: read counts (txt file)
|
| |
|
| Submission date |
Oct 16, 2018 |
| Last update date |
Oct 09, 2019 |
| Contact name |
Tatjana Sauka-Spengler |
| E-mail(s) |
tatjana.sauka-spengler@imm.ox.ac.uk
|
| Organization name |
MRC Weatherall Institute of Molecular Medicine
|
| Department |
University of Oxford
|
| Lab |
Sauka-Spengler lab
|
| Street address |
Radcliffe Department of Medicine
|
| City |
Oxford |
| State/province |
Oxfordshire |
| ZIP/Postal code |
OX3 9DS |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL19787 |
| Series (2) |
| GSE121331 |
Reconstruction of the global neural crest gene regulatory network in vivo [CRISPR_RNA-seq] |
| GSE121527 |
Reconstruction of the global neural crest gene regulatory network in vivo |
|
| Relations |
| BioSample |
SAMN10247163 |
| SRA |
SRX4892461 |
