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| Status |
Public on Sep 15, 2015 |
| Title |
ISC Input |
| Sample type |
SRA |
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| Source name |
Control mouse intestinal stem cells
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| Organism |
Mus musculus |
| Characteristics |
strain: B6;129 mixed background
cell type: Adult intestinal stem cell; proximal 2/3 of intestine
genotype: Cdx2+/+; Lgr5-EGFP-IRES-CREERT2
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| Treatment protocol |
Not injected with tamoxifen.
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| Growth protocol |
Crypts from the proximal 2/3 of mouse small intestines were isolated by scraping off villi and incubating in 5mM EDTA in PBS for 45 minutes at 4°C. Crypts were disaggregated with 4X TrypLE in DMEM for 45 minutes at 37°C and Lgr5-GFP-HI intestinal stem cells were sorted by flow cytometry. Cells were snap-frozen in liquid nitrogen and pooled for subsequent ChIP analysis.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Whole cell lysates were sonicated, but not immunoprecipitated.
The ThruPLEX-FD Prep Kit (Rubicon Genomics) was used to prepare libraries, and 50 bp single-end reads were sequenced on an Illumina Hi-Seq 2000 instrument.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
ISC_Input_DNA
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| Data processing |
Alignment: ChIP-seq:bowtie. RNA-seq: Tophat, version 2.0.9
ChIP-Seq - peak calling: MACS, version 1.4.2, considering input and local background, p<0.0001 cutoff.
RNA-Seq - normalized and differential expression: Cufflinks, version 2.1.1; settings: -b, -u, and default; Cuffmerge was used with custom 3' 1000 bp reference gtf and default settings; Cuffnorm was used with default settings; Cuffdiff was used with -b, -u and default settings.
Genome_build: mm9
Supplementary_files_format_and_content: ChIP-Seq: Bed file for significant peaks, bigwig file for visualizing aligned sequence tags
Supplementary_files_format_and_content: RNA-Seq: There are two processed data matrices that apply to all the samples. First, is the normalized expression table, prepared using Cuffnorm, which contains the normalized Log2(FPKM+1) expression values for all 9 RNA-seq samples. The second is the differential expression table, prepared using Cuffdiff with an FDR of 5%, which contains average FPKM values for villus, control ISC and Cdx2-del ISC, as well as fold change in expression, P-values, and Q-values corrected for multiple hypothesis testing. The third is the custom gtf used in cuffdiff for correction of 3' bias in the RNA-seq samples.
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| Submission date |
Jul 10, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Ramesh Shivdasani |
| E-mail(s) |
ramesh_shivdasani@dfci.harvard.edu
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| Organization name |
Dana Farber Cancer Institute
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| Department |
Medical Oncology
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| Lab |
Shivdasani
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| Street address |
450 Brookline Ave. Dana 720D
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
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| Platform ID |
GPL13112 |
| Series (1) |
| GSE70766 |
Distinct processes and transcriptional targets underlie CDX2 requirements in intestinal stem cells and differentiated villus cells |
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| Relations |
| BioSample |
SAMN03854487 |
| SRA |
SRX1091862 |
| Supplementary data files not provided |
SRA Run Selector |
| Processed data not provided for this record |
| Raw data are available in SRA |
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