Coverage by MeDIP-seq and the HumanMethylation 450K BeadChip of different genomic features.

<p>The different features are described along the bottom axis. 100% coverage is defined as covering all of the elements of a particular type in the human genome. Coverage for MeDIP-seq data (MD-s) (averaged for GM01240 and GM01247) is shown as blue bars and for the HumanMethylation 450K (450K) as red bars. Average percentages covered for each technique for each group of features are given above the bar chart. For MeDIP-seq the region or feature was defined as being covered if any part of the region or feature was covered by or overlapped any part of one or more sequencing reads. The coverage for the MeDIP-seq was consistent between the two samples (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050233#pone.0050233.s005" target="_blank">Table S1</a>), illustrating a high degree of reproducibility for the technique. The coverage shown for the HumanMethylation 450K is reported as the number of features where at least one probe present on the array mapped within the features under consideration i.e. is based on the array design.</p

Concordance of the HumanMethylation 450K (450K) and MeDIP-seq (MD-s) data with bisulfite sequencing (BS-s) data.

<p>The top part of the table gives the concordance of the average beta-values for the 326 probes on the X chromosome from the HumanMethylation 450K (450K) and the methylation score calculated by the MEDIPS software for the MeDIP-seq data (MD-s) to the methylation levels for the bisulfite data (BS-s) from MethTools. The second half of the table contains the concordance for a similar analysis for the HumanMethylation 450K and MeDIP-seq data for all autosomal chromosomes.</p

Comparison of methylation level estimates for the bisulfite sequencing, HumanMethylation 450K and MeDIP-seq data.

<p>Data are shown for the 28 islands (associated with 36 genes) containing CpG sites that overlapped with those interrogated by HumanMethylation 450K array for sample GM01240. Evolutionary strata information is shown to the right of the ideogram of the human X chromosome <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050233#pone.0050233-Ross1" target="_blank">[66]</a>: the blue line represents the S3 stratum; the purple line represents the S2 stratum and the red line the S1 stratum. Both names are given for genes sharing a CpG island separated by “/”. Methylation level estimates for each of the techniques are shown to the right of the gene names in light green (low), green (medium), and dark green (high). Examples of four genes are shown in more detail on the right of the figure. The gene names are highlighted in colour at the top of each panel and in a corresponding colour on the gene list. Data for the bisulfite sequencing (BS-s), HumanMethylation 450K (450K) and MeDIP-seq (MD-s) are shown at the top, center and bottom of each panel, respectively. The genes shown give examples where the three techniques agree in methylation level: low level methylation in the gene ZFX, medium level methylation in the PRPS2 gene, and a high level of methylation in the ACRC gene. Data are also given for the HCFC1/TMEM187 genes, for which different methods show inconsistency in the classified methylation levels. See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050233#pone.0050233.s004" target="_blank">Figure S4</a> for data for sample GM01247.</p

Examples of <i>de novo</i> copy-number variants in offspring.

<p>(<b>A</b>) <i>De novo</i> arisen 67 kb-long deletion on chromosome 6:80596173–80663256 in family T39. Children 1–3 (C010135, C010136 and C010137) have inherited one normal haplotype from both parents. One child (Child 4, C010138) has inherited one normal haplotype from his mother (C010134) and a paternal haplotype with a <i>de novo</i> deletion event in the corresponding region. (<b>B</b>) <i>De novo</i> arisen 167 kb-long duplication on chromosome 2:110175122–110331912 in family T07. The only child (C010026) has inherited one normal haplotype from her mother (C010025) and a paternal haplotype with a <i>de novo</i> intra-chromosomal duplication event in the corresponding region. Coloured arrows show the transmission of specific haplotypes from parents to offspring in a given CNV region. Respective B-allele frequency (BAF, upper panel) and total fluorescent signal intensity (Log R Ratio—LRR, lower panel) plots from Illumina Genome Viewer are shown next to the parents and each child.</p

Examples of unambiguously phased CNV regions involving deletion- and duplication-carrying haplotypes in families.

<p>(<b>A</b>) Inherited 820 kb-long deletion on chromosome 16:15369798–16190572 in family T3. A deletion-carrying haplotype (cn = 0) is inherited from father (C010008) to son (Child 2, C010010). The daughter (Child 1, C010011) has inherited normal haplotypes (cn = 1) from both parents. (<b>B</b>) Inherited 166 kb-long duplication on chromosome 10:47007374–47173619 in family T14. A duplication-carrying haplotype (cn = 2) is inherited from father (C010046) to one son (Child 1, C010049) and daughter (Child 2, C010052). All other children have inherited normal haplotypes (cn = 1) from both parents. Coloured arrows show the transmission of specific haplotypes from parents to offspring in a given CNV region. Respective B-allele frequency (BAF, upper panel) and total fluorescent signal intensity (Log R Ratio—LRR, lower panel) plots from Illumina Genome Viewer are shown next to the parents and each child.</p

Transmission rate of deletion- and duplication-carrying haplotypes in HapMap YRI and EGCUT datasets combined.

<p>*Statistically significant (multiple-testing corrected p-value<0.05) deviations from the expected Mendelian transmission rate of 50%.</p><p>Transmission rate together with the number of transmitted variant-carrying haplotypes and the number of all transmission events (1 event/per locus/per child) for each of the non-overlapping CNV length intervals.</p><p>Transmission rate of deletion- and duplication-carrying haplotypes in HapMap YRI and EGCUT datasets combined.</p

CNV regions in HapMap YRI and EGCUT parents where allelic variability between and within normal and copy number gain-carrying haplotypes can be deterministically differentiated.

<p>In case informative polymorphic genotypes are present <i>between</i> haplotypes in an individual, copy number gain-carrying haplotypes (cn>1) can be deterministically distinguished from the normal single copy haplotypes (cn = 1). Furthermore, these informative genotypes can be used to establish the allelic composition and different allelic copies <i>within</i> copy number gain-carrying haplotypes.</p><p>CNV regions in HapMap YRI and EGCUT parents where allelic variability between and within normal and copy number gain-carrying haplotypes can be deterministically differentiated.</p

Phasing and allelic composition of normal and CNV-carrying haplotypes on parental homologous chromosomes.

<p>A chromosomal region involving copy number variation is denoted with ‘R2’. In the given example, father is the carrier of two normal haplotypes of ‘R2’ on chromosomes P1 and P2 (diploid copy number of ‘R2’, CN = 2), whereas mother has a combination of a duplication-carrying (on M1) and normal (M2) haplotypes (diploid copy number of ‘R2’, CN = 3). Haplotype-informative SNP genotypes in ‘R2’ sequence that can be used for phasing and determining the parental origin (in offspring) of given normal and CNV-carrying haplotypes are given in bold letters and genotypes that are polymorphic <i>between</i> normal or duplication-carrying parental haplotypes are indicated with dashed rectangles. The duplication-carrying haplotype on maternal M1 chromosome is composed of two allelic copies of the sequence ‘R2’ distinguished by genotype variability at position SNP7 (polymorphic SNP variant <i>within</i> the duplication-carrying haplotype), indicated with dotted rectangle.</p

Computational phasing of normal and CNV-carrying haplotypes.

<p>(<b>A</b>) First, CNV and regular two-letter genotypes are collected from the QuantiSNP output for each family member at a locus of interest. (<b>B</b>) Next, markers that have any low-confidence genotype calls or the call could not have been made (‘NC’ genotypes, e.g. marker rs10801575, marked with the red background) and monomorphic markers that are not informative for haplotype phasing in the studied region (e.g. marker rs7517836; marked with the red background) are filtered out. (<b>C</b>) Informative high-confidence genotypes are then phased considering all family members simultaneously and the resulting haplotypes are presented as the result. (<b>D</b>) The family tree of these phased haplotypes can be further visualised for the corresponding CNV region.</p

Detailed description of analysed family-based SNP microarray datasets.

<p>Detailed description of analysed family-based SNP microarray datasets.</p