Method for fragmenting DNA by nick translation

1. A kit for fragmenting double-stranded DNA, comprising:

i. one or more enzymes capable of nick translating activity;

ii. a dNTP having a modified base, wherein the modified base is not fluorescent and cannot participate in a click reaction; and

iii. at least one modification-sensitive nicking endonuclease that are prevented from nicking DNA if its recognition site contains the modified base.

2. The kit of claim 1 , wherein the nicking endonuclease is Nt.CviPII or NciI or both.

3. The kit of claim 1 , wherein the modified base is a modified dCTP or a modified dATP or both.

4. The kit of claim 3 , wherein the dNTP is 5-methylcytosine.

5. The kit of claim 1 , wherein the kit contains a dNTP mix, and the dNTP of (ii) is contained in the dNTP mix.

6. The kit of claim 5 , wherein the dNTP mix comprises: i) 5m dCTP or 7m dATP, ii) 5m dCTP and 7m dATP, iii) 5m dCTP and dCTP, iv) 7m dATP and dATP, v) dUTP, or vi) dUTP and dTTP.

7. The kit of claim 1 , wherein the one or more enzymes capable of nick translating activity is provided by a polymerase having a 5′-3′ exonuclease activity or a strand-displacing polymerase and a flap endonuclease.

8. A composition comprising:

i. one or more enzymes capable of nick translating activity;

ii. a dNTP mix comprising a dNTP having a modified base, wherein the modified base is not fluorescent and cannot participate in a click reaction; and

iii. at least one modification-sensitive nicking endonuclease that is prevented from nicking DNA if its recognition site contains the modified base.

9. The composition of claim 8 , further comprising a double-stranded DNA template that comprises recognition sites for the modification-sensitive nicking endonuclease.

10. The composition of claim 9 , wherein the double-stranded DNA template is genomic DNA, or an amplified form thereof.

11. The composition of claim 8 , further comprising fragments of a double-stranded DNA template, wherein at least some of the fragments comprise the modified base.

12. The composition of claim 11 , wherein the fragments have a desired length selected from a size in the range of 50 bp to 50 kbp.

13. A method for fragmenting DNA comprising:

(a) combining in a reaction mix:

i. one or more polymerases capable of nick translating activity;

ii. a dNTP mix comprising at least one dNTP having a modified base;

iii. at least one modification-sensitive nicking endonuclease that is prevented from nicking DNA if its recognition site contains a modified nucleotide; and

iv. a sample comprising a double-stranded DNA template that comprises recognition sites for the at least one modification-sensitive nicking endonuclease;

(b) incubating the reaction mix for a period of time in order to produce fragments of the double-stranded DNA template, wherein the fragments comprise the at least one modified base; and

(c) ligating adaptors onto the ends of the fragments, amplifying the fragments, and/or sequencing the fragments or an amplification product thereof.

14. The method of claim 13 , wherein the double-stranded DNA template is genomic DNA, or an amplified form thereof.

15. The method of claim 13 , wherein the fragments have a desired length selected from a size in the range of 50 bp to 50 kbp.

16. The method of claim 13 , wherein step (c) comprises ligating adaptors onto the end of the fragments produced in step (b) to produce adaptor-ligated fragments.

17. The method of claim 16 , further comprising amplifying the adaptor-ligated fragments to produce amplification products.

18. The method of claim 13 , wherein step (c) comprises sequencing the fragments produced in step (b), or an amplification product thereof.