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| Status |
Public on Apr 18, 2013 |
| Title |
5hmc-J |
| Sample type |
SRA |
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| Source name |
sperm cells
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| Organism |
Homo sapiens |
| Characteristics |
cell type: sperm
cell phenotype: abnormal sperm cell of fertile men
chemical pulldown target: 5hmc
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Purified genomic DNA should be sonicated into short fragments (200~300 bp) by Covaris DNA shearing with microTUBEs according to manufacturer’s instructions. 5hmC DNA capture was performed using Hydroxymethyl Collect kit. Briefly, 5-hmC labeling reaction were performed in 75-ul solution containing 50mM HEPES buffer(PH7.9), 250 mM MgCl2, 100 µM UDP-6-N3-Glu,and 80 U βGT. The reactions were incubated for 2h at 37℃.After the reaction, The DNA substrates were purified and exchange buffer in H20 via Bio-rad micro bio spin according to the manufacturer’s instructions. The click chemistry was performed with the addition of 150µM biotin into the DNA solution and incubated 2h at 37℃.The DNA samples were then purified by Invitrogen Dynabeads MyOneTM Streptavidin C1 according to manufacturer’s instructions.
5-hmC enriched genomic DNA libraries were generated following the Illumina protocol for “Preparing Samples for CHIP sequencing of DNA”.we used 1ng 5-hmC enriched DNA to initiate the protocol. DNA fragments of 150~300bp were gel purified after the adapter ligation step. PCR-amplified DNA libraries were quantified on an Agilent 2100 Bio analyzer and using quantity PCR.We performed 100bp single end sequencing on Illumina Hiseq2000 to get 5-hmC-enriched DNA fragment sequence.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Pulldown reads were aligned to the hg19 genome assembly using BWA version 0.5.9-r16
Peaks were called using MACS version 1.4.2 with default parameter
Genome_build: hg19
Supplementary_files_format_and_content: MACS-generated peaks file.
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| Submission date |
Apr 17, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Bao-Fa Sun |
| E-mail(s) |
sunbf@big.ac.cn
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| Organization name |
Beijing Institute of Genomics (BIG) of Chinese Academy of Sciences (CAS)
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| Street address |
Da-Tun Road
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| City |
Beijing |
| ZIP/Postal code |
100101 |
| Country |
China |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE46135 |
Epigenome analysis of DNA modification 5-hydroxymethylcytosine in normal, abnormal, and globozoospermia sperm genomes |
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| Relations |
| BioSample |
SAMN02046899 |
| SRA |
SRX268275 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1124795_5hmc_J_peaks.txt.gz |
871.2 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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