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| Status |
Public on Jun 30, 2016 |
| Title |
97_SF/SCL ampulla |
| Sample type |
SRA |
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| Source name |
Ampulla
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| Organism |
Bos indicus |
| Characteristics |
developmental stage: multiparous cows
health status: Healthy
breed: Nelore
tissue: ampulla
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| Treatment protocol |
Animals were pre-synchronized by intramuscular injection of Prostaglandin F2α analog (PGF; 0.5 mg of sodium cloprostenol; Sincrocio, Ouro Fino, Cravinhos, Brazil) and, 7 days later an injection of GnRH agonist (1 µg of buserelin acetate; Sincroforte, Ouro Fino, Cravinhos, Brazil). At this day (D-20) animals received an Estrotect device and estrus detection was performed twice daily from D-20 to D-10. All animals received a new intravaginal P4-releasing device (1g; Sincrogest) on D-10 along with an intramuscular injection of 2 mg estradiol benzoate (Sincrodiol, Ouro Fino). Simultaneously, cows in the LF-LCL received an intramuscular injection of PGF. Additionally, P4 device removal was performed earlier in animals of the LF-LCL group. Specifically, devices were removed 42h and 30 h before the GnRH injection in the LF-LCL and the SF-SCL groups, respectively. All animals received a PGF injection at P4 device removal and a second PGF injection 6h later. Ovulation was induced by an injection of GnRH agonist on D0. On day 4, cows were stunned using captive bolt and euthanized by jugular exsanguination at the University slaughterhouse.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from frozen ampulla and isthmus samples using the AlLrep® DNA/RNA/Protein Mini kit (No. 80004, Qiagen), as recommended by manufacturer’s instructions. cDNA was generated by a routine RNA library preparation TruSeq protocol developed by Illumina Technologies (San Diego, CA) using 1 μg of total RNA as input.
RNA libraries were prepared for sequencing using standard Illumina protocols
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiScanSQ |
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| Data processing |
CASAVA v1.8.2.
seqyClean v1.3.12. (https://bitbucket.org/izhbannikov/seqyclean/get/stable.zip) using 26 Phred quality parameter for maximum average error, and vector and adaptor sequences from the Univec database (https://www.ncbi.nlm.nih.gov/tools/vecscreen/univec/) were use as guide to remove possible contaminants. Only high quality paired-end sequences were kept for further analyses
The reads were mapped with Tophat v.2.0.8 and Bowtie2 v2.1.0 on the masked bovine genome assembly (Bos taurus UMD 3.1, NCBI).
The mapping file was sorted using SAMTools v 0.1.18 and read counts were obtained using the script from HTSeq-count v0.5.4p2 (http://www-huber.embl.de/users/anders/HTSeq/doc/count.html). The differential expression analysis was performed with package DESEq v1.12.1, from R/Bioconductor.
Using the function estimateSizeFactors, the normalized counts were obtained (baseMean values, which are the number of reads divided by the size factor or normalization constant). The standard deviation along the baseMean values was also calculated for each transcript. In order to avoid artifacts caused by low expression profiles and high expression variance, only transcripts that had an average of baseMean > 5 and the mean greater than the standard variation were analyzed.
The method to test for differential expression was the negative binonial distribution, through the nbinomTest function on DESeq. The threshold for evaluating significance was obtained by applying a p-value of 0.05 FDR- Benjamini-Hochberg.
Genome_build: Bos taurus (assembly Bos_taurus_UMD_3.1.1)
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| Submission date |
Feb 05, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Angela Maria Gonella |
| E-mail(s) |
angelamvz@gmail.com
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| Organization name |
Universidade de São Paulo
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| Department |
Departamento de Reprodução Animal
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| Lab |
Laboratório de Fisiologia e Endocrinologia Molecular (LFEM)
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| Street address |
Rua Duque de Caxias Norte, 255
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| City |
Pirassununga |
| State/province |
SP |
| ZIP/Postal code |
13635-900 |
| Country |
Brazil |
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| Platform ID |
GPL19671 |
| Series (2) |
| GSE65679 |
Transcriptome profile form oviduct samples of endocrine manipulated cows (ampulla) |
| GSE65681 |
Transcriptome profile form oviduct samples of endocrine manipulated cows |
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| Relations |
| BioSample |
SAMN02903330 |
| SRA |
SRX793512 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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