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Sample GSM4712035

Query DataSets for GSM4712035

Status

Public on Aug 01, 2021

Title

1_2

Sample type

SRA

Source name

ovarian granulosa cells

Organism

Felis catus

Characteristics

cell type: ovarian granulosa cell
agent: none

Extracted molecule

total RNA

Extraction protocol

Total RNA from untreated and AT-1 treated FOGCs was extracted using TRIzol® reagent (Invitrogen/Thermo Fisher Scientific, Carlsbad, CA, USA) following the manufacturer’s instructions.
To obtain mRNA, an approximately 5 µg sample of total RNA was used to isolate poly (A) mRNA with a poly-T oligo primer attached to magnetic beads (Invitrogen). Sequencing libraries were generated using an NEB Next Ultra™ RNA Library Prep Kit for Illumina (NEB, Ipswich, MA, USA) following the manufacturer’s instructions. To synthesize the first-strand cDNA, purified mRNA was fragmented into small pieces then reverse transcriptase and random hexamer primers were utilized. Synthesis of second-strand cDNA was carried out using deoxynucleoside triphosphates containing DNA polymerase, dUTP, and RNase. End repair was achieved by adding dATP to all free 3’ ends and unique index sequences for the fragments were added with adaptor ligation. A sequencing library was obtained by PCR amplification of the products. The cDNA library was sequenced on an Illumina HiSeq 4000 sequencing platform (Illumina, San Diego, USA ) by Majorbio Bio-pharm Technology Co., Ltd (Shanghai, China).

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 4000

Data processing

The cDNA library was sequenced on an Illumina HiSeq 4000 sequencing platform (Illumina, San Diego, USA ) by Majorbio Bio-pharm Technology Co., Ltd (Shanghai, China).
The raw RNA-seq data was filtered to remove adaptor contamination, low-quality bases, and undetermined bases utilizing Cutadapt software (Maher CA, 2009)
Sequencing quality was verified using FASTQC software including the Q20, Q30, and GC content of the clean data. All downstream analysis was based on clean, high-quality data. The filtered reads were aligned and mapped to the reference genome using Hisat 2.0.
The aligned read files were processed by Cufflinks (Martin M, 2011.), which uses the normalized RNA-seq fragment counts to measure the relative abundances of the transcripts.
The unit of measurement is fragmented per kilobases of exon per million fragments mapped (FPKM). The DEGs were screened using R package edger (Trapnell C, 2009) with Fold change (FC)≥ 1.5 and P < 0.05 (50), which were considered significantly differentially expressed.
Genome_build: Felis_catus_9.0
Supplementary_files_format_and_content: Tab-delimited text files include FPKM values for each sample.

Submission date

Aug 06, 2020

Last update date

Aug 01, 2021

Contact name

Yuli Guo

E-mail(s)

yuliguo@emails.imau.edu.cn

Phone

15849123015

Organization name

Inner Mongolia Agricultural University