This track displays maps of chromatin state in the given cell types as identified by chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-Seq).
In the given cell type, ChIP-Seq begins by using formaldehyde to cross-link histones and other DNA-associated proteins to genomic DNA. The cross-linked chromatin is subsequently extracted, mechanically sheared, and immunoprecipitated using specific antibodies. After reversal of cross-links, the immunoprecipitated DNA is sequenced and mapped to the human reference genome. The relative enrichment of each antibody-target (epitope) across the genome is inferred from the density of mapped fragments.
Measurements were made in neuronal and non-neuronal nuclei collected from prefrontal cortex (PFC) of 11 individuals ranging in age from 0.5 to 69 years.
Human prefrontal cortex samples used in this study were obtained from the Brain and Tissue Bank for Developmental Disorders, University of Maryland and a brain bank at the University of California, Irvine. Nuclei extraction, chromatin immunoprecipitation and sequencing were carried out as described in Cheung I, et al. (2010). Sequencing was performed on an Illumina Genome Analyzer (GA II).
The length of sequence reads was 36 bp. Reads were mapped using Bowtie (version 0.11.3, Langmead B, Trapnell C, Pop M, Salzberg SL (2009)) allowing up to one mismatch to map all sequence reads to the gender appropriate human genome hg18, and 67-87% of the reads in the neuronal samples mapped to one unique location in the genome. The mapped reads were analyzed using the MACS software package (version 1.3.5, Zhang Y, et al. (2008)) with bw = 230 bp, as defined experimentally by PCR, tSize = 36 bp, and other parameters set at default.
A subset of the peaks that were positioned more than 10 kb from annotated genes were experimentally tested for RNA expression in PFC neurons by qRT-PCR and in situ hybridization to verify that H3K4me3 mapping can serve as a guide to uncover unannotated novel and cell-specific transcripts in the brain.
Chromatin immunoprecipitation experiments were carried out by Iris Cheung, Yan Jiang and Schahram Akbarian; analyses were performed by Hennady Shulha, Jie Wang and Zhiping Weng at the University of Massachusetts Medical School.
Cheung I, et al. Developmental regulation and individual differences of neuronal H3K4me3 epigenomes in the prefrontal cortex. Proc. Natl. Acad. Sci. USA (in press for May 2010 publication).
Langmead B, Trapnell C, Pop M, Salzberg S. Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol. 2009;10:R25.
Zhang Y, Liu T, Meyer CA, Eeckhoute J, Johnson DS, Bernstein BE, Nussbaum C, Myers RM, Brown M, Li W, et al. 2008. Model-based Analysis of ChIP-Seq (MACS). Genome Biol 9: R137