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      Coverage recommendations for methylation analysis by whole genome bisulfite sequencing

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          Abstract

          Whole genome bisulfite sequencing (WGBS) allows genome-wide DNA methylation profiling but the associated high sequencing costs continue to limit its widespread application. We utilized several high coverage reference data sets to experimentally determine minimal sequencing requirements. Here, we present data derived recommendations for minimum sequencing depth for WGBS libraries, highlight what is gained with increasing coverage and discuss the trade off between sequencing depth and number of assayed replicates.

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          Stability and flexibility of epigenetic gene regulation in mammalian development.

          Wolf Reik (2007)
          During development, cells start in a pluripotent state, from which they can differentiate into many cell types, and progressively develop a narrower potential. Their gene-expression programmes become more defined, restricted and, potentially, 'locked in'. Pluripotent stem cells express genes that encode a set of core transcription factors, while genes that are required later in development are repressed by histone marks, which confer short-term, and therefore flexible, epigenetic silencing. By contrast, the methylation of DNA confers long-term epigenetic silencing of particular sequences--transposons, imprinted genes and pluripotency-associated genes--in somatic cells. Long-term silencing can be reprogrammed by demethylation of DNA, and this process might involve DNA repair. It is not known whether any of the epigenetic marks has a primary role in determining cell and lineage commitment during development.
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            Transcriptional and epigenetic dynamics during specification of human embryonic stem cells.

            Differentiation of human embryonic stem cells (hESCs) provides a unique opportunity to study the regulatory mechanisms that facilitate cellular transitions in a human context. To that end, we performed comprehensive transcriptional and epigenetic profiling of populations derived through directed differentiation of hESCs representing each of the three embryonic germ layers. Integration of whole-genome bisulfite sequencing, chromatin immunoprecipitation sequencing, and RNA sequencing reveals unique events associated with specification toward each lineage. Lineage-specific dynamic alterations in DNA methylation and H3K4me1 are evident at putative distal regulatory elements that are frequently bound by pluripotency factors in the undifferentiated hESCs. In addition, we identified germ-layer-specific H3K27me3 enrichment at sites exhibiting high DNA methylation in the undifferentiated state. A better understanding of these initial specification events will facilitate identification of deficiencies in current approaches, leading to more faithful differentiation strategies as well as providing insights into the rewiring of human regulatory programs during cellular transitions. Copyright © 2013 Elsevier Inc. All rights reserved.
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              Towards sound epistemological foundations of statistical methods for high-dimensional biology.

              A sound epistemological foundation for biological inquiry comes, in part, from application of valid statistical procedures. This tenet is widely appreciated by scientists studying the new realm of high-dimensional biology, or 'omic' research, which involves multiplicity at unprecedented scales. Many papers aimed at the high-dimensional biology community describe the development or application of statistical techniques. The validity of many of these is questionable, and a shared understanding about the epistemological foundations of the statistical methods themselves seems to be lacking. Here we offer a framework in which the epistemological foundation of proposed statistical methods can be evaluated.
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                Author and article information

                Journal
                101215604
                32338
                Nat Methods
                Nat. Methods
                Nature methods
                1548-7091
                1548-7105
                10 October 2014
                02 November 2014
                March 2015
                01 September 2015
                : 12
                : 3
                : 230-232
                Affiliations
                [1 ]Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA
                [2 ]Harvard Stem Cell Institute, Cambridge, Massachusetts 02138, USA
                [3 ]Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, Massachusetts 02138, USA
                [4 ]McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins School of Medicine, Baltimore, Maryland 21205, USA
                [5 ]Department of Biostatistics, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland 21205, USA
                [6 ]Molecular Pathology Unit, Massachusetts General Hospital, Charlestown, Massachusetts 02129, USA
                [7 ]Center for Cancer Research, Massachusetts General Hospital, Charlestown, Massachusetts 02129, USA
                [8 ]Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA
                Author notes
                Article
                NIHMS634114
                10.1038/nmeth.3152
                4344394
                25362363
                1057ec52-084c-448c-b86a-9258e70b67c8
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                Life sciences
                Life sciences

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