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Updated: Dec 16, 2025

3' End Sequencing Library Preparation with A-seq2

3' End Sequencing Library Preparation with A-seq2

Published on: October 10, 2017

Tet-Assisted Bisulfite Sequencing (TAB-seq).

Miao Yu¹, Dali Han^2,3, Gary C Hon⁴

¹Ludwig Institute for Cancer Research, La Jolla, CA, 92093, USA.

Methods in Molecular Biology (Clifton, N.J.)

|December 11, 2017

Summary

This summary is machine-generated.

Tet-Assistant Bisulfite Sequencing (TAB-Seq) offers single-base resolution for detecting 5-Hydroxymethylcytosine (5hmC). This method quantifies 5hmC abundance, overcoming limitations of previous low-resolution techniques.

Keywords:

5-Hydroxymethylcytosine Bisulfite sequencing DNA methylation Glycosylation TAB-Seq

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Last Updated: Dec 16, 2025

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Enhanced Reduced Representation Bisulfite Sequencing for Assessment of DNA Methylation at Base Pair Resolution

Enhanced Reduced Representation Bisulfite Sequencing for Assessment of DNA Methylation at Base Pair Resolution

Published on: February 24, 2015

Area of Science:

Epigenetics and Genomics
Molecular Biology Techniques

Background:

5-Hydroxymethylcytosine (5hmC) is a crucial epigenetic modification in mammalian cells, derived from 5-methylcytosine (5mC) via Ten-eleven translocation (TET) protein oxidation.
Existing methods for profiling 5hmC have limitations in resolution (~100 bp) and lack quantitative site-specific abundance data.
Understanding 5hmC's role in gene regulation necessitates high-resolution, quantitative detection methods.

Purpose of the Study:

To present a detailed protocol for Tet-Assistant Bisulfite Sequencing (TAB-Seq).
To enable single-base resolution detection and quantification of 5hmC abundance.
To provide a method that overcomes the resolution and quantification limitations of current 5hmC profiling techniques.

Main Methods:

Genomic DNA is treated with β-glucosyltransferase (βGT) to convert 5hmC to 5-glucosylhydroxymethylcytosine (5hmC).
Subsequent treatment with recombinant mouse Ten-eleven translocation 1 (mTet1) converts 5-methylcytosine (5mC) to 5-carboxylcytosine (5caC).
The modified DNA is then subjected to bisulfite treatment for locus-specific detection or whole-genome sequencing.

Main Results:

The developed TAB-Seq protocol achieves single-base resolution for 5hmC detection.
TAB-Seq provides quantitative information on the abundance of 5hmC at each specific genomic site.
The protocol is adaptable for both targeted analysis of specific loci and comprehensive whole-genome analysis.

Conclusions:

Tet-Assistant Bisulfite Sequencing (TAB-Seq) is a powerful new method for high-resolution, quantitative 5hmC profiling.
TAB-Seq significantly advances the ability to study the functional roles of 5hmC in epigenetic regulation.
This protocol offers a valuable tool for epigenetic research in mammalian cells and tissues.