|
| Status |
Public on May 05, 2017 |
| Title |
Lgr5 negative subpopulation from P4 (CD24+CD29hi), replicate 3 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
MaSC
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57/B6
genotype/variation: lgr5-egfp-ires-creert2
mg condition: Virgin
lgr5 status: Negative
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cell preparation protocol: Single cell suspensions were prepared from the mammary glands of virgin mice using mechanical and enzymatic dissociation as detail described in Tiede BJ et al., 2009, PLoS ONE 4(11): e8035. doi:10.1371/journal.pone.0008035
FACS sorting protocol: Various subpopulations of mammary epithelial cells were sorted using flow cytometry based on combination of a set of surface markers and the expression of EGFP. The marker profiles are detailed in Tiede BJ et al., 2009, PLoS ONE 4(11): e8035. doi:10.1371/journal.pone.0008035 and Plaks 2013 (PMID: 23352663). FACS gating parameters are detailed in Plaks 2013 (PMID: 23352663). Specifically, Lin-CD24+CD29hiGFP+ is Lgr5 positive P4 and Lin-CD24+CD29hiGFP- is Lgr5 negative P4.
Total RNA were extracted using RNAeasy mini-prep kit from Qiagen
|
| Label |
Cy5
|
| Label protocol |
The testing RNA samples and universal mouse reference RNA (Agilent) were labeled with CTP-cy5 and CTP-cy3, respectively, Using Agilent Quick Amp Labeling Kit.
|
| |
|
| Channel 2 |
| Source name |
Universal Mouse Reference RNA
|
| Organism |
Mus musculus |
| Characteristics |
cell lines: Mix of 11 mouse cell lines
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cell preparation protocol: Single cell suspensions were prepared from the mammary glands of virgin mice using mechanical and enzymatic dissociation as detail described in Tiede BJ et al., 2009, PLoS ONE 4(11): e8035. doi:10.1371/journal.pone.0008035
FACS sorting protocol: Various subpopulations of mammary epithelial cells were sorted using flow cytometry based on combination of a set of surface markers and the expression of EGFP. The marker profiles are detailed in Tiede BJ et al., 2009, PLoS ONE 4(11): e8035. doi:10.1371/journal.pone.0008035 and Plaks 2013 (PMID: 23352663). FACS gating parameters are detailed in Plaks 2013 (PMID: 23352663). Specifically, Lin-CD24+CD29hiGFP+ is Lgr5 positive P4 and Lin-CD24+CD29hiGFP- is Lgr5 negative P4.
Total RNA were extracted using RNAeasy mini-prep kit from Qiagen
|
| Label |
Cy3
|
| Label protocol |
The testing RNA samples and universal mouse reference RNA (Agilent) were labeled with CTP-cy5 and CTP-cy3, respectively, Using Agilent Quick Amp Labeling Kit.
|
| |
|
| |
| Hybridization protocol |
Each cy5 labeled testing sample was mixed with equal amount cy3-labeled reference sample and fragmented and blocked (Agilent Gene Expression Hybridization Kit), and samples were applied to microarrays enclosed in Agilent SureHyb-enabled hybridization chambers. After hybridization, slides were washed sequential
|
| Scan protocol |
Scanned on an Agilent G2505C scanner.
Images were quantified using Agilent Feature Extraction Software (version 11.0).
|
| Description |
See Agilent Cat#740100 for product details
|
| Data processing |
Agilent Feature Extraction Software (v 11.0) was used for background subtraction and linear Lowess normalization.
|
| |
|
| Submission date |
Aug 15, 2016 |
| Last update date |
Feb 29, 2024 |
| Contact name |
Yong Wei |
| E-mail(s) |
[email protected]
|
| Organization name |
Princeton University
|
| Department |
Molecular Biology
|
| Lab |
Kang
|
| Street address |
LTL227 Wathington Road
|
| City |
Plainsboro |
| State/province |
NJ |
| ZIP/Postal code |
08536 |
| Country |
USA |
| |
|
| Platform ID |
GPL10787 |
| Series (2) |
| GSE85633 |
Transcriptome characterization of the mouse Lgr5+_MaSCs |
| GSE85808 |
miR-199a-LCOR axis regulation of mammary gland stem cell and breast cancer stem cell transcriptional programs |
|
