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Sample GSM5510406

Query DataSets for GSM5510406

Status

Public on Nov 10, 2021

Title

follicular_granulosa_cell_layers-SWF_RNA-seq_replicate_2

Sample type

SRA

Source name

granulosa cells

Organism

Gallus gallus

Characteristics

cell type: granulosa cells
breed: Luhua hens
developmental stage: prehierarchical follicles
age: 31 weeks
follicle growth stage: SWF

Extracted molecule

total RNA

Extraction protocol

total RNA was extracted from each sample using RNAiso Plus reagent (TaKaRa, Otsu, Shiga, Japan) following the manufacturer’s instructions.
Strand Synthesis Reaction Buffer(5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase(RNase H-). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3’ ends of DNA fragments, NEB Next Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 250~300 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). Then 3 μl USER Enzyme (NEB, USA) was used with size-selected, adaptor-ligated cDNA at 37°C for 15 m in followed by 5min at 95 °C before PCR. Then PCR was performed with Phusion High -Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system.
We estimated the integrity and quality of total RNA using a Bioanalyzer 2100 system (Agilent Technologies, Palo Alto, CA, USA) with an RNA 6000 Nano kit. Subsequently, 30 strand-specific libraries (total of 10 stages with three biological replicates in each stage) were generated after depleting rRNA by the Ribo-Zero™ Gold Kit (Illumina, San Diego, CA, USA) and then sequenced on the Illumina HiSeq X Ten platform (Illumina Inc., San Diego, CA, USA) with a paired-end sequencing length of 150 bp (PE150) at Novogene Corporation (Beijing, China).

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

DNBSEQ-T7

Description

bulk.RNAseq.all_counts.filt.0.5tpm.more.than.3num.xlsx
TPM-submission.xlsx
SWF-2

Data processing

TPM-submission.xlsx: bulk-RNAseq reads were aligned to the galGal6 using kallisto version 0.44.0 ,then tpm and counts matrixs were generated using R version 3.6.1.
bulk.RNAseq.all_counts.filt.0.5tpm.more.than.3num.xlsx: bulk-RNAseq reads were aligned to the galGal6 using kallisto version 0.44.0 ,then tpm and counts matrixs were generated using R version 3.6.1.
SWF.filtered_feature_bc_matrix.h5: single cell reads were aligned to the galGal6 using cellranger version 6.0.1
F1.filtered_feature_bc_matrix.h5: single cell reads were aligned to the galGal6 using cellranger version 6.0.1
POF.filtered_feature_bc_matrix.h5: single cell reads were aligned to the galGal6 using cellranger version 6.0.1
ATAC-SWF_peaks.xls: ATAC-seq reads were algned to the galGal6 using bowtie2 version 2.2.6, mitochondrial alignments and PCR duplicates were filtered using removeChrom and Picard version 1.126 and peaks were called using macs2 version 2.1.1.
ATAC-F1_peaks.xls: ATAC-seq reads were algned to the galGal6 using bowtie2 version 2.2.6, mitochondrial alignments and PCR duplicates were filtered using removeChrom and Picard version 1.126 and peaks were called using macs2 version 2.1.1.
ATAC-POF_peaks.xls: ATAC-seq reads were algned to the galGal6 using bowtie2 version 2.2.6, mitochondrial alignments and PCR duplicates were filtered using removeChrom and Picard version 1.126 and peaks were called using macs2 version 2.1.1.
IP-SWF_AllEnhancers.table.txt: chip-seq reads were aligned to the galGal6 using BWA version 0.7.15, data were filtered using samtools version 1.3.1 and super-enhancers were defined using ROSE algorithms.
IP-F1_AllEnhancers.table.txt: chip-seq reads were aligned to the galGal6 using BWA version 0.7.15, data were filtered using samtools version 1.3.1 and super-enhancers were defined using ROSE algorithms.
IP-POF_AllEnhancers.table.txt: chip-seq reads were aligned to the galGal6 using BWA version 0.7.15, data were filtered using samtools version 1.3.1 and super-enhancers were defined using ROSE algorithms.
F1.POF.silico.bulk.DEG.result.by.DEseq2.txt: single cell reads were aligned to the galGal6 using cellranger version 6.0.1, silico bulk-RNAseq counts matrixs were extracted using Python 2.7 and DEGs were generated using DEseq2 algorithms.
SWF.F1.silico.bulk.DEG.result.by.DEseq2.txt: single cell reads were aligned to the galGal6 using cellranger version 6.0.1, silico bulk-RNAseq counts matrixs were extracted using Python 2.7 and DEGs were generated using DEseq2 algorithms.
SWF.POF.silico.bulk.DEG.result.by.DEseq2.txt: single cell reads were aligned to the galGal6 using cellranger version 6.0.1, silico bulk-RNAseq counts matrixs were extracted using Python 2.7 and DEGs were generated using DEseq2 algorithms.
Genome_build: galGal6
Supplementary_files_format_and_content: bulk-RNAseq (xlsx files)
Supplementary_files_format_and_content: single cell RNAseq (matrix.h5 files)
Supplementary_files_format_and_content: ATAC-seq (peaks.xls files)
Supplementary_files_format_and_content: chip-seq (AllEnhancers.table.txt files)
Supplementary_files_format_and_content: single cell RNAseq (silico.bulk.DEG.result.by.DEseq2.txt files)

Submission date

Aug 10, 2021

Last update date

Nov 10, 2021

Contact name

hua kui

E-mail(s)

2020102003@stu.sicau.edu.cn

Organization name

Sichuan Agricultural University

Street address

No 211 Huimin Road