|
| Status |
Public on Feb 19, 2013 |
| Title |
Stallion sperm transcriptome 2 |
| Sample type |
SRA |
| |
|
| Source name |
Stallion Spermatozoa
|
| Organism |
Equus caballus |
| Characteristics |
fertility stutus: Normal fertile male
tissue: Sperm
|
| Treatment protocol |
Fresh sperm ejaculates from reproductively normal stallions were collected using an artificial vagina. The ejaculates were first evaluated for sperm concentration, motility characteristics and morphological features, followed by purification from somatic cells and immature sperm by EquiPure™ (Nidacon International, Sweden) discontinuous gradient centrifugation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from sperm with TRIzol reagent (Invitrogen). Total RNA from the sperm of reproductively normal stallions was used for next generation sequencing (NGS) on the ABI SOLiD platform. THE TOTAL 500 ng of total RNA was directly used for SOLiD single-end RNA sequencing fragment library construction according to the ABI protocol
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
AB SOLiD System 3.0 |
| |
|
| Data processing |
The 50 bp single-end SOLiD raw reads were directly aligned with the horse reference sequence EcuCab2 using ABI aligner software (NovoalignCS version 1.00.09, novocraft.com) which uses multiple indexes in the reference genome, identifies candidate alignment locations for each primary read, and allows completion of the alignment.
The single highest-scoring alignment for each raw read was mapped. Alignments in repetitive sequences were discarded by removing reads with multiple similarly scoring alignments (non-unique matches).
Sequence alignment and alignment clustering to define expressed loci and perform linear normalization across the two sperm RNA samples was carried out with a software package EXP (Cofactor Genomics).
Gene expression level or average coverage (AC) was calculated by normalizing each sample to the fewest reads and directly comparing different loci. Expression level of a transcript was estimated from the number of reads that mapped to that transcript.
The variability present in sequencing depths in different samples was taken care of by the use of two biological replicates. Sequencing depth at each locus and differences in gene expression (AC) between the two sperm samples were calculated using log (base2) ratio, thus showing the association between the two samples.
Genome_build: EcuCab2
Supplementary_files_format_and_content: tab-delimited text files include chromosomal location of 19,257 RNA Seq tag combining two biological replicate with their expression value (Average Coverage), Average coverage≥1
|
| |
|
| Submission date |
Jun 14, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Pranab Jyoti Das |
| E-mail(s) |
pjdas@cvm.tamu.edu
|
| Phone |
979-458-0520
|
| Fax |
979-845-9972
|
| URL |
http://vetmed.tamu.edu/labs/cytogenics-genomics
|
| Organization name |
Texas A&M University
|
| Department |
Department of Veterinary Integrative BioSciences
|
| Lab |
Molecular Molecular Cytogenetics and Genomics Laboratory
|
| Street address |
Room 314B Bldg. 1197
|
| City |
College Station |
| State/province |
TX |
| ZIP/Postal code |
77843 |
| Country |
USA |
| |
|
| Platform ID |
GPL15696 |
| Series (1) |
| GSE38725 |
Stallion sperm transcriptome as revealed by microarray analysis and RNA sequencing |
|
| Relations |
| SRA |
SRX154656 |
| BioSample |
SAMN01054173 |
