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| Status |
Public on Mar 07, 2017 |
| Title |
Input_H3K27me3_Ctrl1_ChIPSeq |
| Sample type |
SRA |
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| Source name |
B-LCL_Ctrl1
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| Organism |
Homo sapiens |
| Characteristics |
cell type: cell line
genotype: DNMT3Bwt/wt
chip antibody: none
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| Growth protocol |
ICF and control B-LCL were coltured in RPMI-1640 (SIGMA) supplemented with 10% fetal bovine serum.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei. Histone-DNA and DNMT3B-DNA complexes were isolated with specific antibodies. Immunoprecipitated complexes were recovered with protein A sepharose (Pharmacia), washed with low and high salt buffers, reverse-crosslinked, and purified.
DNMT3B and H3K36me3 libraries were prepared according to Illumina's instructions accompanying the Nextera DNA Sample Kit. H3K27me3 and H3K4me3 libraries were prepared according to SOLiD protocol (Applied Biosystem).
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
AB SOLiD 4 System |
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| Description |
Input DNA
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| Data processing |
DNMT3B and H3K36me3 sequence reads were generated by Illumina's HiSeq2000 platform. H3K27me3 and H3K4me3 reads were sequenced using the SOLiD platform (Applied Biosystem).
Quality of the short reads of 50 bp were analysed and high quality reads were selected. They were aligned to the human reference genome (hg19) by using Bowtie v. 1.1.1. The parameters used for Bowtie were –a, –m3, –-best, and –-strata.
Data were filtered by uniquely aligned reads and a single read for each starting position was considered.
DNMT3B peak calling was performed using MACS v.1.4.2. The sample enrichments were normalized for cell line specific input. MACS parameter settings were: --bw 200, -p 1e-4 and -m 8,30. Moreover, we detected large peaks using EDD (12), setting: log_ratio_bin_size 2Kb, -g 5 and -n 20000. Confidence intervals were calculated using the normal approximation method for binomial proportions. The SICER v.1.1 (13) peak-finding algorithm was used to identify the H3K4me3-, H3K27me3- and H3K36me3-enriched sites throughout the genome. For all these histone marks we selected peaks with a false discovery rate (FDR) of 1e-5. We used as window size and gap different values accordingly with the profile of analyzed histone marks. In particular, for H3K4me3 we used window size 200 bp and gap size 200 bp, for H3K27me3 500 bp and 1000 bp, while for H3K36me3 800bo and 4000bp respectively.
Genome_build: hg19
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| Submission date |
Mar 06, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Miriam Gagliardi |
| E-mail(s) |
miriam_gagliardi@psych.mpg.de
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| Organization name |
Max Planck Institute of Psychiatry
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| Street address |
Kraepelinstrasse 2-10
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| City |
Munich |
| State/province |
Bayern |
| ZIP/Postal code |
80804 |
| Country |
Germany |
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| Platform ID |
GPL13393 |
| Series (2) |
| GSE95746 |
ICF-specific DNMT3B dysfunction interferes with intragenic regulation of mRNA transcription and alternative splicing (ChIP-Seq) |
| GSE95747 |
ICF-specific DNMT3B dysfunction interferes with intragenic regulation of mRNA transcription and alternative splicing |
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| Relations |
| BioSample |
SAMN06481957 |
| SRA |
SRX2614789 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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