|
| Status |
Public on Apr 21, 2020 |
| Title |
control2dpi |
| Sample type |
SRA |
| |
|
| Source name |
small intestinal crypts
|
| Organism |
Mus musculus |
| Characteristics |
tissue: small intestinal crypts
strain/genotype: Tg(Vil1-cre/ERT2); Ctnnb1-wt/flox
days post cre induction: 2
background strain: C57BL/6
|
| Treatment protocol |
To induce Cre-ERT2-mediated recombination, tamoxifen (Sigma, 80 mg/kg) was injected intraperitoneally on two consecutive days. Mice were sacrificed 48 hours after the first tamoxifen injection. Alternatively, mice were injected on 4 consecutive days and sacrificed 96 hours after the first injection.
|
| Growth protocol |
All animals were kept on a C57BL/6 background. Mice were 8-12 weeks old at the time of treatments and cell isolations. Mice of both sexes were used in all experiments and littermates were used as controls.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Control (n=3) and D164A (n=3) mice were sacrificed 0, 2 or 4d post injection, crypts were isolated as described (Sato and Clevers, 2013). Crypt fractions 3 and 4 were pooled and dissociated to single cells by incubating them with 5 mL pre-warmed TripLE Express Enzyme (ThermoFischer) in a 37 °C water bath for 5 min with frequent agitation. TripLE was inactivated with 50% FBS in Advanced DMEM/F12 (ThermoFischer). Single cells were pelleted at 180 g for 3 min and resuspended in 2 mL cold Intesticult (Stemcell Technologies Inc). Clumps were dissolved by pipetting up and down with a 1mL pipet before filtering twice through a 40 micron filter. Cell viability and number were quantified in automatically with a Countess (Thermo Scientific).
DropSeq workflow was performed as described (Macosko et al, 2015) on a Nadia Instrument (Dolomite Bio). Sequencing was performed on the Illumina NovaSeq6000 with an SP Reagent Kit.
single cell RNASeq
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
FastQC
zUMIs pipeline
Genome_build: Mus_musculus.GRCm38.95
Supplementary_files_format_and_content: count matrix
|
| |
|
| Submission date |
Apr 20, 2020 |
| Last update date |
Apr 22, 2020 |
| Contact name |
Tomas Valenta |
| E-mail(s) |
tomas.valenta@mls.uzh.ch
|
| Organization name |
University of Zurich
|
| Department |
Institute of Molecular Life Sciences
|
| Street address |
Winterthurerstrasse 190
|
| City |
Zurich |
| ZIP/Postal code |
CH-8057 Zurich |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL24247 |
| Series (2) |
| GSE148942 |
Discrete regulation of β-catenin-mediated transcription governs identity of intestinal epithelial stem cells [scRNA-Seq] |
| GSE148943 |
Discrete regulation of β-catenin-mediated transcription governs identity of intestinal epithelial stem cells |
|
| Relations |
| BioSample |
SAMN14645510 |
| SRA |
SRX8142722 |
