 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jun 01, 2024 |
| Title |
fetal-Rumen, 1 (atac-seq) |
| Sample type |
SRA |
| |
|
| Source name |
Rumen
|
| Organism |
Bos taurus |
| Characteristics |
tissue: Rumen
Sex: male
age: 90 days fetus
genotype: WT
|
| Treatment protocol |
For pluripotent stem cells: The bovine embryonic stem cell line bESCs-F7 was established using the CTFR system. Two treatment methods were employed for bESCs-F7 using MM-102: one involved treating bESCs-F7 with a high concentration (50 μM) of MM-102 for 7 days, followed by normal culturing for 5 passages before characterization, labeled as bESCs-F7-102 (50). The other involved long-term treatment with a low concentration (5 μM) of MM-102, followed by characterization after 5 passages of culturing, labeled as bESCs-F7-102 (5). Bovine induced pluripotent stem cells and the bEPSCs-B18 cell line were established using the LCDM system. And bEPSCs-AGS was using a 500mL system, including 485mL of basic mTeSR1 medium, 5.0 mL of 100×Penicillin-Streptomycin Solution, 0.1 mM 2-mercaptoethanol, 1 μM GSK3β inhibitor CHIR99021, 0.3 μM Lck/Src inhibitor WH-4-023, 5 μM Tankyrase inhibitor XAV939, 5 μM classic WNT signaling pathway inhibitor IWR-1, 50 μg/mL Vitamin C, 10 ng/mL Leukemia Inhibitory Factor (LIF), and 20.0 ng/mL Activin A for culturing bovine Embryonic Pluripotent Stem Cells.
|
| Growth protocol |
For tissues, we collected fetal tissues from four healthy pregnant Holstein cows at a slaughterhouse. After the slaughter, we extracted fetal tissues to examine their normal development and measured the crown-rump length (CRL) to estimate gestational age. The gender was identified based on the position of the gonads within the abdominal cavity, anatomical structures, and relationships with adjacent organs (mesonephros and/or kidneys). Two male and two female fetuses were identified (13-16 cm CRL), nine different tissues were collected, including the forebrain, hindbrain, heart, liver, lungs, skeletal muscle (hind limb), kidneys, rumen, and testes.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA degradation and contamination was monitored on agarose gels. DNA purity was checked using the NanoPhotometer® spectrophotometer (IMPLEN, CA, USA). DNA concentration was measured using Qubit® DNA Assay Kit in Qubit® 2.0 Flurometer (Life Technologies, CA, USA).
A total amount of 5.2 microgram genomic DNA spiked with 26 ng lambda DNA were fragmented by sonication to 200-300bp with Covaris S220, followed by end repair and adenylation. Cytosine-methylated barcodes were ligated to sonicated DNA as per manufacturer’s instructions. Then these DNA fragments were treated twice with bisulfite using EZ DNA Methylation-GoldTM Kit (Zymo Research), before the resulting single-strand DNA fragments were PCR amplificated using KAPA HiFi HotStart Uracil + ReadyMix (2X). Library concentration was quantified by Qubit® 2.0 Flurometer (Life Technologies, CA, USA) and quantitative PCR, and the insert size was assayed on Agilent Bioanalyzer 2100 system.
|
| |
|
| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
First of all, we use FastQC (fastqc_v0.11.5) to perform basic statistics on the quality of the raw reads. Then, those reads sequences produced by the Illumina pipleline in FASTQ format were pre-processed through Trimmomatic (Trimmomatic-0.36) software use the parameter (SLIDINGWINDOW: 4:15 ; LEADING:3, TRAILING:3 ; ILLUMINACLIP: adapter.fa: 2: 30: 10 ; MINLEN:36) The remaining reads that passed all the filtering steps was counted as clean reads and all subsequent analyses were based on this. At last, we use FastQC to perform basic statistics on the quality of the cleandata reads.
Bismark software (version 0.16.3; Krueger F, 2011) was used to perform alignments of bisulfite-treated reads to a reference genome (-X 700 --dovetail). The reference genome was firstly transformed into bisulfite-converted version (C-to-T and G-to-A converted) and then indexed using bowtie2 (Langmead B, 2012). Sequence reads were also transformed into fully bisulfite-converted versions (C-to-T and G-to-A converted) before they are aligned to similarly converted versions of the genome in a directional manner. Sequence reads that produce a unique best alignment from the two alignment processes (original top and bottom strand) are then compared to the normal genomic sequence and the methylation state of all cytosine positions in the read is inferred. The same reads that aligned to the same regions of genome were regarded as duplicated ones. The sequencing depth and coverage were summarized using deduplicated reads.
The results of methylation extractor (bismark_methylation_extractor, --no_overlap) were transformed into bigWig format for visualization using IGV browser. The sodium bisulfite non-coversion rate was calculated as the percentage of cytosine sequenced at cytosine reference positions in the lambda genome.
Differentially methylated regions (DMRs) were identified using the DSS software (Hao Feng HaoWu, 2014；Hao Wu，2015；Yongseok Park Hao Wu,2016 ) , The core of DSS is a new dispersion shrinkage method for estimating the dispersion parameter from Gamma-Poisson or Beta-Binomial distributions.
Assembly: UCSC-BT9
Supplementary files format and content: narrowPeak
|
| |
|
| Submission date |
Apr 18, 2024 |
| Last update date |
Jun 01, 2024 |
| Contact name |
jing wang |
| E-mail(s) |
nnlrl@mail.imu.edu.cn
|
| Organization name |
Inner Mongolia University
|
| Street address |
Zhaojun Road
|
| City |
Hohhot |
| State/province |
Nei Mongol |
| ZIP/Postal code |
010000 |
| Country |
China |
| |
|
| Platform ID |
GPL26012 |
| Series (2) |
| GSE264333 |
Epigenetic Basis for the Establishment of Ruminal Tissue-Specific Functions (ATAC-seq) |
| GSE264346 |
Epigenetic Basis for the Establishment of Ruminal Tissue-Specific Functions |
|
| Relations |
| BioSample |
SAMN41008391 |
| SRA |
SRX24302397 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8216264_fetal_rumen_REP1.mLb.clN_peaks.narrowPeak.gz |
47.6 Kb |
(ftp)(http) |
NARROWPEAK |
SRA Run Selector |
| Raw data are available in SRA |
|
|
|
|
 |
