|
| Status |
Public on Aug 09, 2016 |
| Title |
wt Rep3 |
| Sample type |
SRA |
| |
|
| Source name |
wt
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain: Sprague-Dawley
gender: male
age: 6 weeks old
tissue: Hypothalamus
genotype: +/y (WT)
|
| Treatment protocol |
Not applicable
|
| Growth protocol |
Rats were maintained on a 12 h light:12 h dark cycle with standard rat chow (Pico Lab Rodent Diet, #5053, Purina USA) and water ad libitum. Animals were generated by mating female Sprague-Dawley rats lacking one copy of functional MeCP2 (Mecp2ZFN/+) with wild-type male Sprague-Dawley rats to obtain male rats completely lacking MeCP2 (Mecp2ZFN/y), and wild-type littermate male rats.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Rats were euthanized under anesthesia according to approved IACUC protocols. Hypothalami were rapidly dissected from brain tissue samples on ice and immediately snap frozen in liquid nitrogen. RNA extraction (N=4 biological replicates of each genotype, male Mecp2ZFN/y and male wild-type littermate rats, 6 weeks of life) was performed using TRizol reagent (ThermoFisher Scientific) and further purified with on-column DNase treatment using the RNeasy Mini Kit (Qiagen) following the manufacturers’ instructions.
Ten µg of RNA was prepared for paired-end RNA-Sequencing using Illumina HiSeq 2500. The Baylor College of Medicine Genomic and RNA Profiling Core first conducted Sample Quality checks of the RNA using the NanoDrop ND-1000 spectrophotometer and Agilent Bioanalyzer 2100 using the Nano chip. The Illumina EpiCentre Ribo-Zero rRNA Removal Kit was used to reduce the ribosomal RNA content. The rRNA-reduced sample was then carried into the Illumina TruSeq RNA library preparation protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
SP34044
|
| Data processing |
Illumina HiSeq 2500 software used for basecalling
Raw reads were first groomed by removing 10bp from the 5'-ends with random base concentration, then mapped to Rattus norvegicus genome (Ensembl RGSC3.4) using TopHat using TopHat v1.4.1 (-r 400 –p 8).
Expression level for each gene was estimated using Python program HTSeq (Anders et al., Bioinformatics 2014). Specifically htseq-count function of HTSeq accumulated the number of aligned reads that falls under the exons of the gene (union of all the exons of the gene). This read counts obtained are analogous to the expression level of the gene.
Using the obtained read counts, differential gene analyses were carried out using the DESeq package in the R environment.
Genome_build: Ensembl RGSC3.4
Supplementary_files_format_and_content: Text files inclue read counts value for each gene; individual file for each sample.
|
| |
|
| Submission date |
Jun 14, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Rodney C Samaco |
| E-mail(s) |
rodney.samaco@bcm.edu
|
| Organization name |
Baylor College of Medicine
|
| Department |
Molecular and Human Genetics
|
| Street address |
1250 Moursund St.
|
| City |
Houston |
| State/province |
TX |
| ZIP/Postal code |
77030 |
| Country |
USA |
| |
|
| Platform ID |
GPL18694 |
| Series (1) |
| GSE83323 |
Loss of MeCP2 in the rat models regression, impaired sociability and transcriptional deficits of Rett syndrome |
|
| Relations |
| BioSample |
SAMN05242663 |
| SRA |
SRX1842841 |
