#

Genome-Wide DNA Hydroxymethylation Analysis

Simply send in your samples and receive publication-ready data!

Zymo Research's services for DNA hydroxymethylation analysis offer unparalleled sensitivity and coverage of 5-hydroxymethylcytosine (5-hmC). The majority of previous DNA methylation (5-mC) studies utilized bisulfite conversion (i.e., bisulfite treatment), which cannot distinguish between 5-mC and 5-hmC marks. This is an important consideration because the functional consequences of these two modifications are very different. Therefore, to determine what contribution was made by 5-hmC in past DNA methylation studies, these experiments should be revisited using tools that can accurately detect both modifications.

Services for both single-base resolution and enrichment platforms for specific detection of 5-hmC in DNA are available from Zymo Research. Both platforms combine unique whole-genome library preparation with the latest Next-Gen sequencing technology to ensure high coverage and sensitivity.


Reduced Representation Hydroxymethylation Profiling (RRHP)

The first and only service available for genome-wide strand-specific analysis of 5-hmC at single-nucleotide resolution.

  • Accurate and cost effective
  • Extremely sensitive and quantitative
  • Compatible with low DNA input and FFPE/LCM/FACS sorted samples
  • Use together with Methyl-MiniSeq to profile 5-mC & 5-hmC in the same sample
  • No harsh chemical conversion or treatment steps
  • Simultaneously detect SNPs in the analysis

5-hmC CapSeq / JBP1-Seq

Genome-wide detection of 5-hmC "peaks" that indicate regions containing 5-hmC in DNA.

  • Accurate and cost effective
  • More specific and sensitive than antibody-based or biotin-streptavidin pull-down methods
  • Detect 5-hmC in any sequence motif, without bias
  • Simultaneously detect SNPs in the analysis

Published Methods

JBP1-seq: A fast and efficient method for genome-wide profiling of 5-hmC

Zymo Research scientists have developed an affinity based approach for genome wide 5-hmC profiling that combines the benefits of an improved recombinant JBP1 protein with Nextera-based NGS library construction. Recombinant JBP1 is biotinylated in vivo and conjugated to magnetic beads via biotin–streptavidin interactions. These modifications allow efficient and consistent pull-down of ß-glucosyl-5-hydroxymethylcytosine (ß-glu-5-hmC), and sequence-ready libraries can be generated within 4.5 h from DNA inputs as low as 50 ng. Comparison of technical duplicates and validations with alternative platforms demonstrate the method to be highly reproducible and reliable, thus providing a fast, efficient, and cost-effective method for accurate 5hmC genome-wide profiling.

RRHP: a tag-based approach for 5-hydroxymethylcytosine mapping at single-site Resolution

Current methods for genomic mapping of 5-hydroxymethylcytosine (5hmC) have been limited by either costly sequencing depth, high DNA input, or lack of single-base resolution. Zymo Research scientists present an approach called Reduced Representation 5-Hydroxymethylcytosine Profiling (RRHP) to map 5hmC sites at single-base resolution by exploiting the use of beta-glucosyltransferase to inhibit enzymatic digestion at the junction where adapters are ligated to a genomic library. By doing so, only library fragments presenting glucosylated 5hmC residues at the junction are sequenced. RRHP can detect sites with low 5hmC abundance, and when combined with RRBS data, 5-methylcytosine and 5-hydroxymethylcytosine can be compared at a specific site.