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| Status |
Public on May 10, 2019 |
| Title |
Serum Reponse of CF1 primary MEFs , E13 |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
The goal of this study was to find the longitudinal transcriptional response of Mouse Embryonic Fibroblast (MEF) primary cells during the progression of a cell cycle. Using RNASeq method (single-end 50-bp chemistry on Illumina Hi-seq 2500 instrument in high-throughput mode), we measured the transcriptional response for around two cell-cycles. Using the fold-change (with respect to average response before serum addition at t = 0) time-series data, we first identified, without using a priori knowledge, the duration and timing of cell cycle phases using a change-point detection algorithm. Next, using Least Absolute Shrinkage and Selection Operator (LASSO) and Estimation Stability with Cross Validation (ES-CV), we were able to, without any prior biological knowledge, extract information on the phase-specific causal interaction of cell cycle genes, as well as temporal interdependencies of biological mechanisms through a complete cell cycle.
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| Overall design |
MEF cells were serum starved for 36 hours (Serum Starved Conditions - 0.5% FCS, DMEM High Glucose, L-glutamine, Penicillin Streptomycin, fungizone, non-essential amino acids) prior to adding serum at t = 0 hr. At t = 0, FCS was added to 20% level by volume to stimulate cell growth. Cells were extracted using Trizol RNA isolation protocol with scraping. Total RNA was isolated using Zymo Research Direct-zol 96 RNA (R2054) kit. TruSeq Stranded mRNA HT Kit (RS-122-2101) was used for library preparation. mRNA was sequenced (single-end 50-bp) using Illumina HiSeq 2500 in high throughput mode, with 12 samples per lane. Data processing steps are: 1) Demultiplexing using Casava [configureBclToFastq.pl --mismatches 1], 2) Mapping/Alignment using RNA-STAR [STAR --genomeDir mm9 --sjdbGTFfile mouse_splice.sjdb --outSammode FULL --outFilterType BySJout], and 3) Normalization and Quantification using Homer [analyzeRepeats.pl rna mm9 -count exons -strand both].
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| Contributor(s) |
Pao G, Ke E, Ogawa J, Tonnu N, Verma IM, Masnadi-Shirazi M, Maurya MR, Subramaniam S |
| Citation(s) |
31142274 |
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| Submission date |
Dec 18, 2018 |
| Last update date |
Jun 21, 2019 |
| Contact name |
Shankar Subramaniam |
| Organization name |
University of California, San Diego
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| Department |
Bioengineering
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| Street address |
9500 Gilman Dr MC 0412
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| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92093-0412 |
| Country |
USA |
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| Platforms (1) |
| GPL17021 |
Illumina HiSeq 2500 (Mus musculus) |
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| Samples (96)
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| Relations |
| BioProject |
PRJNA510577 |
| SRA |
SRP173789 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE124024_nonredun_Filter.txt.gz |
4.7 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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