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Status |
Public on Feb 11, 2019 |
Title |
DNA methylation_monkey_SPA |
Sample type |
SRA |
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Source name |
spermatogonia
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Organism |
Macaca mulatta |
Characteristics |
cell type: spermatogonia
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Treatment protocol |
For the gonocyte or spermatogonia, we separated them from the testis tissue of half-year-old rhesus monkeys. Only spermatogonia could be observed in the seminiferous tubules of testis tissue at this age, through hematoxylin-eosin staining and stage-specific antibody staining (PGP9.5). Testis cells were also collected through two step enzyme digestion. The first step used an enzyme solution in DMEM and the second step used an enzyme solution in 0.25% trypsin EDTA, both containing 1mg/ml collagenase, 1mg/ml DNaseⅠand 2.5 mg/ml hyaluronidase (Cat# H3884, Sigma).
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Extracted molecule |
genomic DNA |
Extraction protocol |
The DNA extraction follows Stem-seq protocol The STEM-seq libraries were generated using the protocol as described previously(Zhang et al., 2018). Briefly, cells were lysed in lysis buffer (10mM Tris-Hcl PH 7.5, 10mM NaCl,0.5% NP-40, protease K (Roche)) for 3h at 55°C and heat-inactivated for 1h at 72°C. Then we added Lamda DNA as a control to calculate bisulfite conversion rate. The DNA mixture was digested by dsDNA fragmentase (NEB) and followed by bisulfite conversion using EpiTect Fast Bisulfite Conversion Kit (Qiagen). Finally, the purified single strand converted DNA is tailed by poly C, followed by extension with a biotinylated poly G containing primer. The extension product is ligated to an adaptor followed by PCR amplification for sequencing library preparation. All libraries were sequenced on Illumina HiSeq XTen according to the manufacturer’s instruction. TELP
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Library strategy |
Bisulfite-Seq |
Library source |
genomic |
Library selection |
RANDOM |
Instrument model |
HiSeq X Ten |
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Data processing |
Basecalls performed using CASAVA version 1.8 For Hi-C sapples: sisHi-C sequencing reads were mapped, processed and iteratively corrected using HiC-Pro, a pipeline developed by Servant et al. Briefly, the read pairs were mapped to the mm9 reference genome in a two-step approach with bowtie2. Then the invalid read pairs including dangling end, self-circle ligation and duplicates were discarded. The genome was divided into bins of specific length to generate the contact maps. For global detection of contacts, a 100Kb bin size was used and a 40Kb bin size was used for examination of local domain level contacts. The raw contact counts are normalized with iterative correction. For methylation samples: STEM-seq reads were aligned to the rheMac2 genome using BSseeker2.0.8, and methylation value in bedGraph files were counted by the number of reads falling into 200bp bin in the genome. For RNA-seq samples: SMART-seq2 reads were aligned to the rheMac2 or mm9 genome assembly using tophat2 version 2.0.11, then replicates were merged together, and transcript abundance (FPKM) were calculated based on Refseq annotation using cufflinks version 2.0.2. Genome_build: rheMac2, mm9 Supplementary_files_format_and_content: The bedgraph files contains the values counted by the number of reads falling into 200bp bin in the genome. The gene fpkm txt file contains FPKM value for all samples. with number for each bin. While the validpair txt file indicates the paired interaction reads for each sample.
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Submission date |
Jan 18, 2018 |
Last update date |
Feb 11, 2019 |
Contact name |
Wei Xie |
E-mail(s) |
xiewei121@tsinghua.edu.cn
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Organization name |
Tsinghua University
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Street address |
Zhongguancun north street
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City |
Beijing |
ZIP/Postal code |
100084 |
Country |
China |
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Platform ID |
GPL24522 |
Series (1) |
GSE109344 |
Reprogramming of meiotic chromatin architecture during primate spermatogenesis |
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Relations |
BioSample |
SAMN08375227 |
SRA |
SRX3591090 |
Supplementary file |
Size |
Download |
File type/resource |
GSM2940109_monkey_spa_mCG.bedgraph.gz |
44.4 Mb |
(ftp)(http) |
BEDGRAPH |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
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