|
| Status |
Public on Jun 04, 2018 |
| Title |
CD49fHi Lgr5Hi biological replicate 3 |
| Sample type |
RNA |
| |
|
| Source name |
CD49fHi Lgr5Hi mammary cells from Lgr5-IRES-GFP female embryos isolated at E14
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Embryonic mouse mammary gland FACS purified cells
|
| Treatment protocol |
Pregnant females bearing Lgr5-IRES-GFP embryos were sacrificed at 14 days of gestation and E14 embryos were collected and epidermis + mammary gland were digested in Collagenase+hyaluronidase. Single cells were stained with the appropriate antibodies and FACS sorted.
|
| Growth protocol |
Mice were maintained in a certified animal house according to European guidelines
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA isolation and cDNA amplification was performed as described by Gonzalez-Roca et al. (2010).In brief, 813 or 2000 cells were sorted into lysis buffer and RNA was purified using magnetic beads (RNAClean XP beads, Agencourt).RNA was reverse transcribed and cDNA was amplified using Whole Transcriptome Amplification chemistry (WTA2, Sigma Aldrich). For monitoring amplification, SYBR Green was added to the reaction; it was stopped after 25 cycles when SYBR Green signal reached a plateau. cDNA was purified using Genelute PCR Clean-Up kit (Sigma Aldrich).
|
| Label |
Biotin
|
| Label protocol |
cDNA was labeled using GeneChip Mapping 250K Nsp Assay Kit (Affymetrix; catalog # 900766), according to manufacturer's instructions.
|
| |
|
| Hybridization protocol |
Affymetrix MG-430 PM array strips were hybridized with 8 mg of labeled cDNA, washed, stained and scanned according to the protocol described in GeneAtlas 3’IVT express kit User Manual.
|
| Scan protocol |
Arrays were scanned with GeneAtlasImaging station scanner (Affymetrix, Santa Clara, CA)
|
| Description |
Gene expression of 813 CD49fHi Lgr5Hi mammary cells from Lgr5-IRES-GFP female embryos isolated at E14, biological replicate 3
|
| Data processing |
All the results were normalized using the RMA normalization algorithm using R-bioconductor affy package with standard parameters. Cross experiment normalization was further performed to eliminate the batch effect using non-parametric empirical Bayes frameworks for adjusting data implemented in ComBat function of the Surrogate Variable Analysis package (SVA) in R-bioconductor for the two different microarrays used here and then between this experiment and the previously published data under the accession number GSE69290.
|
| |
|
| Submission date |
Jan 17, 2018 |
| Last update date |
Jun 04, 2018 |
| Contact name |
Alexandra Van Keymeulen |
| Organization name |
Université Libre de Bruxelles
|
| Department |
IRIBHM
|
| Lab |
Blanpain
|
| Street address |
808, route de Lennik CP602
|
| City |
Bruxelles |
| ZIP/Postal code |
1070 |
| Country |
Belgium |
| |
|
| Platform ID |
GPL11180 |
| Series (2) |
| GSE109306 |
Early lineage segregation of multipotent embryonic mammary gland progenitors [expression] |
| GSE109711 |
Early lineage segregation of multipotent embryonic mammary gland progenitors |
|
