Compositions comprising glucosylated hydroxymethylated bases
The present invention provides for novel methods for regulating and detecting the cytosine methylation status of DNA. The invention is based upon identification of a novel and surprising catalytic activity for the family of TET proteins, namely TET1, TET2, TET3, and CXXC4. The novel activity is related to the enzymes being capable of converting the cytosine nucleotide 5-methylcytosine into 5-hydroxymethylcytosine by hydroxylation.
1. An isolated nucleic acid from an extracellular fluid sample, wherein a hydroxymethylated cytosine of said isolated nucleic acid is glucosylated.
2. The nucleic acid of claim 1 , wherein said isolated nucleic acid comprises genomic DNA.
3. The nucleic acid of claim 1 , wherein said glucosylated hydroxymethylated cytosine is a glucosylated 5-hydroxymethlycytosine.
4. The nucleic acid of claim 1 , wherein a glucose of said glucosylated hydroxymethylated cytosine is labeled.
5. The nucleic acid of claim 1 , further comprising a guanine (G), an adenine (A), a thymine (T), a uracil (U), and a cytosine (C).
6. A double-stranded DNA, wherein a strand of said double-stranded DNA comprises said isolated nucleic acid of claim 1 .
7. A composition comprising said isolated nucleic acid of claim 1 and a nucleic acid sequence complementary to at least a portion of said isolated nucleic acid.
8. The composition of claim 7 , wherein said nucleic acid sequence, that is complementary to at least a portion of said isolated nucleic acid, is a population of nucleic acid sequences.
9. A composition comprising said isolated nucleic acid of claim 1 and a DNA polymerase.
10. A composition comprising said isolated nucleic acid of claim 1 and a solid support.
11. The composition of claim 10 , wherein said glucosylated hydroxymethylated cytosine is associated with said solid support.
12. The composition of claim 10 , wherein a moiety is associated with said glucosylated hydroxymethylated cytosine.
13. The composition of claim 12 , wherein said moiety is covalently coupled to said glucosylated hydroxymethylated cytosine.
14. The composition of claim 12 , wherein said moiety comprises a biotinyl group.
15. A method comprising: contacting said isolated nucleic acid of claim 1 with a moiety, wherein said moiety comprises a biotinyl group or a fluorophore.
16. A method comprising: isolating said isolated nucleic acid of claim 1 from a sample by affinity purification.
17. A method comprising: sequencing said isolated nucleic acid of claim 1 .
18. A composition comprising: (a) a glucosylated nucleic acid comprising a glucosylated 5-hydroxymethylcytosine; and (b) an isolated and purified dioxygenase enzyme or fragment thereof.
19. The composition of claim 18 , further comprising a co-factor.
20. The composition of claim 19 , wherein said co-factor comprises iron.
21. The composition of claim 18 , wherein said glucosylated nucleic acid is labeled.
22. A composition comprising: (a) glucosylated nucleic acid; and (b) an isolated and purified dioxygenase enzyme or said fragment thereof that is a TET family enzyme or fragment thereof.
23. The composition of claim 22 , further comprising a co-factor.
24. The composition of claim 23 , wherein said co-factor comprises iron.
25. The composition of claim 22 , wherein said glucosylated nucleic acid is labeled.