Compositions, methods and related uses for cleaving modified DNA
View Patent ↗Compositions, methods and related uses are provided relating to cleaving modified DNA. For example, a set of DNA fragments obtainable by enzymatic cleavage of a large DNA is described where at least 50% are similarly sized and have a centrally positioned modified nucleotide. In addition, an enzyme preparation is provided that includes one or more enzymes that recognize a modified nucleotide in a DNA and cleave the DNA at a site that is at a non-random distance from the modified nucleotide. The one or more enzymes are further characterized by an N-terminal conserved domain with greater than 90% amino acid sequence homology to WXD(X) 10 YXGD. The related uses include creating a methylome, methods of purifying DNA fragments containing a modified nucleotide and diagnostic applications.
1. A composition comprising a plurality of similar sized double-stranded fragments, the fragments further characterized by:
(a) staggered ends; and
(b) a centrally positioned methylated or hydroxymethylated cytosine proximal to a guanine;
wherein the fragments are obtainable by in vitro enzymatic cleavage of a large DNA, the large DNA comprising genomic DNA.
2. The composition according to claim 1 , wherein plurality of fragments are isolated.
3. The composition according to claim 1 , wherein the large DNA is at least 100 nucleotides in length.
4. The composition according to claim 1 , wherein the large DNA is a mammalian genomic DNA.
5. The composition according to claim 1 , wherein the large DNA is human genomic DNA.
6. The composition according to claim 1 , wherein the fragments are less than 60 nucleotides in size.
7. The composition according to claim 6 , wherein the fragments have a similar size in the range of 28-36 nucleotides.
8. The composition according to claim 1 , wherein the centrally positioned methylated or hydroxymethylated cytosine is located within 30 nucleotides from one end of the fragments.
9. The composition according to claim 1 , wherein the centrally positioned methylated or hydroxymethylated cytosine and guanine are in a CpG dinucleotide.
10. The composition according to claim 1 , wherein the centrally positioned methylated or hydroxymethylated cytosine and guanine are in a CNG trinucleotide.