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Direct, programmable detection of epigenetic DNA cytosine modifications using TAL effectors

  • US 9,879,315 B2
  • Filed: 06/26/2014
  • Issued: 01/30/2018
  • Est. Priority Date: 06/26/2013
  • Status: Active Grant
First Claim
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1. A method for the differentiation of a 5-hydroxymethyl modification of a cytosine residue of interest in a nucleic acid molecule from(i) a 5-methyl modification of said cytosine residue of interest or(ii) an unmodified cytosine residue of interest,said method comprising the steps of:

  • (a) providing a transcription-activator-like effector (TALE) protein, said TALE protein being capable of binding in a sequence- and locus- specific manner to a region of said nucleic acid molecule that includes said cytosine residue of interest, wherein a repeat variable diresidue (RVD) of said TALE protein whose position corresponds to the position of the cytosine residue of interest is a size reduced RVD; and

    (b) determining whether said TALE protein binds to said region bydetecting a 5-hydroxymethyl modification of said cytosine residue of interest, wherein(i) said modification is present when said TALE protein does not bind to said region or does bind to said region with a strongly reduced affinity, and(ii) said modification is absent when said TALE protein binds to said region,said method allowing for the direct detection of a 5-hydroxymethyl modification of said cytosine residue of interest in said nucleic acid molecule.

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