|
| Status |
Public on Oct 11, 2016 |
| Title |
KO male E3.5, repeat 2 [Rlim KO] |
| Sample type |
SRA |
| |
|
| Source name |
Whole embryo
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
genotype: Rlim KO
Stage: E3.5
gender: male
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Whole embryos were dissected at the indicated timepoints and the correct stage was verified under the binocular. Single embryos/trophoblast cells were placed in 10ul TCL Buffer (Qiagen) supplemented with 1% BME, and snap frozen.
RNA libraries were prepared for sequencing using standard Illumina protocols. A total of 187 samples were distributed on two 96-well plates (plus 5 mock wells) and thawed at RT for 10 minutes prior to RNA purification using Ampure RNA beads (Beckmann-Coulter). RNA samples were re-suspended in solution containing 3’ RT primer and dNTPs. Reverse transcription was performed with SSII (Life Technologies), whose terminal transferase activity allows incorporation of a PCR binding site at the 3’ end of the cDNA using a template-switching oligonucleotide (custom synthesized from Exiqon) as a template. Subsequently, cDNA was amplified using 12 cycles of PCR, followed by tagmentation with Nextera kit (Illumina). Final libraries were amplified by 12 cycles of PCR and sequenced on a NextSeq 500.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
The bcl2fastq 2.16.0.10 software package was used to convert base call (BCL) files generated on the NextSeq system to fastq files
Sequenced reads were aligned to the mouse genome (mm10) using TopHat (version 2.0.12) with default setting except set parameter read-mismatches to 2
HTSeq (version 0.6.1p1) was used to count the features after the alignment
Bioconductor packages edgeR (version 3.10.0 ) and ChIPpeakAnno (version 3.2.0) for transcriptome quantification, differential gene expression analysis and annotation.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include the raw count values from HTSeq for each sample
|
| |
|
| Submission date |
Jun 17, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Ingolf Bach |
| E-mail(s) |
ingolf.bach@umassmed.edu
|
| Organization name |
University of Massachusetts Medical School
|
| Department |
Molecular, Cell and Cancer Biology
|
| Street address |
364 Plantation St
|
| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
| |
|
| Platform ID |
GPL19057 |
| Series (2) |
| GSE69970 |
Transcriptome of mouse preimplantation development [Rlim KO] |
| GSE71442 |
Transcriptome of mouse preimplantation development |
|
| Relations |
| BioSample |
SAMN03780137 |
| SRA |
SRX1063852 |
