METHODS OF EPIGENETIC ANALYSIS
The present invention provides for methods of epigenetic analysis. In some cases, the methods may include obtaining a sample comprising a nucleic acid sequence. In some cases, the nucleic acid sequence may comprise one or more epigenetic marks. The methods may include performing a sequencing. The methods may include distinguishing a hydroxymethylated base from a methylated base.
1 .- 10 . (canceled)
11 . A method, comprising:
contacting a polynucleotide with an enzyme or a catalytically active fragment thereof that performs an oxidation on an epigenetic modification to form a contacted polynucleotide; and
sequencing the contacted polynucleotide.
12 . The method of claim 11 , wherein the enzyme or the catalytically active fragment performs a conversion selected from the group consisting of:
oxidizing 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC);
oxidizing 5mC to an acid beyond hydroxymethylation;
oxidizing 5mC to an aldehyde beyond hydroxymethylation;
oxidizing 5hmC to an acid beyond hydroxymethylation;
oxidizing 5hmC to an aldehyde beyond hydroxymethylation; and any combination thereof.
13 . The method of claim 11 , further comprising isolating the polynucleotide from a sample.
14 . The method of claim 11 , wherein the polynucleotide is isolated from an extracellular fluid.
15 . The method of claim 11 , wherein the enzyme or the catalytically active fragment thereof is isolated prior to contacting.
16 . The method of claim 11 , wherein the polynucleotide is contacted with the enzyme.
17 . The method of claim 11 , wherein the polynucleotide is contacted with the catalytically active fragment of the enzyme.
18 . The method of claim 11 , wherein the enzyme or the catalytically active fragment thereof comprises a catalytically active TET family enzyme or a functional fragment thereof.
19 . The method of claim 11 , wherein the sequencing comprises determining a methylation state of the contacted polynucleotide.
20 . The method of claim 11 , wherein the sequencing comprises an imaging.
21 . The method of claim 11 , wherein the polynucleotide comprises 5mC and wherein the contacting converts at least in part the 5mC to 5hmC.
22 . The method of claim 11 , wherein the polynucleotide comprises 5mC and wherein the contacting converts at least in part the 5mC to an acid beyond hydroxymethylation.
23 . The method of claim 11 , wherein the polynucleotide comprises 5mC and wherein the contacting converts at least in part the 5mC to an aldehyde beyond hydroxymethylation.
24 . The method of claim 11 , wherein the polynucleotide comprises 5hmC and wherein the contacting converts at least in part the 5hmC to an acid beyond hydroxymethylation.
25 . The method of claim 11 , wherein the polynucleotide comprises 5hmC and wherein the contacting converts at least in part the 5hmC to an aldehyde beyond hydroxymethylation.
26 . The method of claim 11 , wherein the enzyme comprises a TET1 enzyme or a catalytically active fragment thereof.
27 . The method of claim 11 , wherein the enzyme comprises a TET2 enzyme or a catalytically active fragment thereof.
28 . The method of claim 11 , wherein the enzyme comprises a TET3 enzyme or a catalytically active fragment thereof.
29 . The method of claim 11 , wherein the enzyme comprises a CXXC4 enzyme or a catalytically active fragment thereof.
30 . The method of claim 11 , further comprising identifying a 5mC.
31 . The method of claim 11 , further comprising identifying a 5hmC.