Epigenetic Analysis

Genome-wide methylation arrays – Illumina Methylation 450K and EPIC arrays (Human)

Arrays offer a reliable, cost-effective way of analysing multiple targets on a large scale. The Genome Centre has offered Illumina methylation arrays since their original golden gate assay. We have now phased over from their Meth450 array to the new EPIC array.

Illumina have steadily increased the content on their Infinium-based mathylation arrays from 27,000 targets, through 450,000 targets, to their latsest version – the EPIC array – which covers 850,000 targets. Each of the array formats covers 99 per cent of RefSeq promoters and also targets miRNA and cancer-specific sites giving cost-effective and comprehensive analysis of the methylome. The EPIC also includes FANTOM5 and ENCODE identified methylations sites. Details of the original array design can be found in Bibikova et al 2012.

The Genome Centre service requires 1µg of genomic DNA per sample and can utilise degraded DNA from FFPE samples (see Thirwell et al 2011). Submitted samples are Quaity Controlled (QC) prior to bisulphite treatment followed by on array analysis.

Data can be provided either as raw image, intensity or text files following QC and basic analysis with Illumina's GenomeStudio.

We can also survey methylated cytosines at the genome-wide level using MeDip and Bisulphite Next Generation Sequencing or individual loci scales with pyrosequencing.

Illumina Methylation Array Properties
Array type	CpG sites assayed	Samples per array	Content	DNA required	Ref
Meth 450	450,000+	12	99 per cent RefSeq genes	1µg	Bibikova et al 2012 datasheet_human_methylation450 [PDF 740KB] Meth450 data sheet
EPIC	850,000+	8	99 per cent RefSeq genes, ENCODE and FANTOM5 targets	1µg	datasheet_human_methylationEPIC [PDF 1,141KB] EPIC data sheet

To discuss projects or for costings, please contact Dr Charles Mein on 020 7882 2055 or by email at c.a.mein@qmul.ac.uk.

Single site methyl cytosine analysis

In addition to discovery or screening analysis of methylated-cytosine, individual or clustered CpG sites can be accurately interrogated using pyrophosphate sequencing on the pyrosequencing PSQ96 platform (see Bogdarina et al 2007).

Genomic DNA is bisulphite-treated modifying unmethylated cytosines to uracils whilst leaving methylated cytosines intact. Modified DNA is then amplified using PCR followed by short read sequencing on the pyrosequencer PSQ96, usually a few 10s of base pairs, to assay up to six consecutive CpG sites. Intensity differences in base incorporation at the CpG site indicate the proportion of methylation at any one site.

Our services

We are able to support this application from initial consultation through to analysis. A typical project would follow the following process:

1. Consultation discussion of assay design and constraints
2. Design with Pyromark Assay Design v2.0.1.15 software
3. Assay optimisation with unlabelled PCR primers
4. Synthesis of successful assays
5. Sample QC
6. Bisulphite treatment
7. Pyrosequencing
8. Analysis and data delivery with PyroQ-CpG 1.0.9 software

Sample requirements are:

• A minimum of 1µg genomic DNA
  A minimum of 50bp of DNA sequence around CpG site of interest