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Sample GSM2948737 Query DataSets for GSM2948737
Status Public on Jun 04, 2018
Title AW1-CD1-LC1.1
Sample type SRA
 
Source name mammary gland
Organism Mus musculus
Characteristics tissue: mammary gland
gender: female
mouse strain: CD1
cell surface markers: CD45-CD31-CD140a-CD24+CD29low
Treatment protocol Targeting Confetti, tdTomato or ΔNp63-IRES-GFP expression in the MG – For clonal lineage tracing, K14rtTA/TetO-Cre/Rosa-Confetti embryos were induced at E13 by intravenous (IV) injection in the tail vein of the pregnant mother with 1µg/g of Doxycycline (diluted in sterile PBS, Sigma) and sacrificed 2 days later (E15) or 5 days after delivery (P5). K5CreER/Rosa-Confetti newborn pups (P1) were induced by intraperitoneal (IP) injection of 50µg of Tamoxifen (diluted in sunflower seed oil, Sigma) and sacrificed 21 days later. For lineage tracing at saturation, K14rtTA/TetO-Cre/Rosa-tdTomato or K14rtTA/TetO-Cre/Rosa-ΔNp63-IRES-GFP embryos were induced at E13 by IV injection in the tail vein of the pregnant mother with 15µg/g of Doxycycline (diluted in sterile PBS) and sacrificed 5 days after delivery (P5). K8rtTA/TetO-Cre/Rosa-Confetti female mice were induced at 4w old with Doxycyclin treatment during 7 days consisting in the combination of Doxycylin 10g/kg in diet (Bio-Serv), 2g/l in drinking water (AG Scientific) and 3 intraperitoneal injections of 2 mg in 200µl PBS.
Growth protocol Mice – Lgr5-EGFP-IRES-CreER18 and Rosa-tdTomato63 mice were obtained from the Jackson Laboratory. Rosa-Confetti15 mice were provided by H. Clevers; K14rtTA transgenic mice64 were provided by Elaine Fuchs; TetO-Cre mice65 were provided by Andreas Nagy; Rosa26-ΔNp63-IRES-GFP mice57 were provided by Wim Declercq. The generation of K5CreER and of K8rtTA were previously described9, 66. All experimental mice used in this study were females of mixed genetic backgrounds. No statistical methods were used to predetermine sample size. The experiments were not randomized. The investigators were not blinded to allocation during experiments and outcome assessment. Mice colonies were maintained in a certified animal facility in accordance with European guidelines. The experiments were approved by the local ethical committee (CEBEA).
Extracted molecule total RNA
Extraction protocol RNAseq and analysis of bulk samples40000 LCs and 5000 BCs were isolated by FACS as described above and collected into kit lysis buffer. RNA was extracted using absolutely RNA nanoprep kit (Stratagene).
RNA quality was checked using a Bioanalyzer 2100 (Agilent technologies). Indexed cDNA libraries were obtained using the Ovation Solo RNA-Seq System (NuGen) following manufacturer recommendation. The multiplexed libraries (18 pM) were loaded on flow cells and sequences were produced using a NovaSeq 6000 S2 Reagent Kit (200 cycles) from a NovaSeq 6000 System (Illumina).
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
 
Description CP20M-Wuidart
Data processing Approximately 19 million of paired-end reads per sample were mapped against the mouse reference genome (GRCm38/mm10) using STAR software to generate read alignments for each sample. Annotations Mus_musculus.GRCm38.87.gtf were obtained from ftp.Ensembl.org. After transcripts assembling, gene level counts were obtained using HTSeq. Fold change of mean gene expression for the duplicates were used to calculate the level of differential gene expression. Heatmap of the 500 most variable genes across the 8 samples and corresponding clustering dendrogram were drawn with heatmap.2 function of the R package gplots (citation R). Euclidian distance with complete linkage agglomeration method was used for clustering.
Genome_build: grcm38
Supplementary_files_format_and_content: CP20M-Wuidart (count normalized by 20M mapped reads
 
Submission date Jan 26, 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 GPL24247
Series (2)
GSE109707 Early lineage segregation of multipotent embryonic mammary gland progenitors [RNA-seq I]
GSE109711 Early lineage segregation of multipotent embryonic mammary gland progenitors
Relations
BioSample SAMN08400238
SRA SRX3602389

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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