Methods of characterizing small nucleic acid molecules
The invention generally relates to methods of characterizing small nucleic acid molecules, such as plasmids or nucleic acid molecules obtained from a virus. In certain embodiments, the invention provides a method of characterizing a small circular nucleic acid molecule, the method including performing rolling circle amplification on a small circular nucleic acid molecule, thereby producing a concatamer, and generating an optical map of the concatamer, thereby characterizing the small circular nucleic acid molecule.
1. A method of characterizing a nucleic acid molecule, the method comprising:
performing rolling circle amplification on a nucleic acid molecule of 1,000 base pairs or fewer, thereby producing a concatamer; and
generating an optical map of the concatamer, thereby characterizing the nucleic acid molecule.
2. The method according to claim 1 , wherein prior to said performing step, the method further comprises circularizing a nucleic acid molecule of 1,000 base pairs or fewer.
3. The method according to claim 1 , wherein the concatamer comprises at least five internal restriction sites.
4. The method according to claim 1 , wherein the nucleic acid molecule is DNA or RNA.
5. The method according to claim 1 , wherein the nucleic acid molecule is a plasmid.
6. The method according to claim 1 , wherein the nucleic acid molecule is obtained from a virus.
7. The method according to claim 1 , wherein the nucleic acid molecule is single-stranded.
8. The method according to claim 1 , wherein the nucleic acid molecule is double-stranded.
9. A method of characterizing a nucleic acid molecule, the method comprising:
circularizing a nucleic acid molecule of 1,000 base pairs or fewer to produce a circular nucleic acid molecule;
performing rolling circle amplification on the circular nucleic acid molecule, thereby producing a concatamer; and
generating an optical map of the concatamer, thereby characterizing the nucleic acid molecule.
10. The method according to claim 9 , wherein the concatamer comprises at least five internal restriction sites.
11. The method according to claim 9 , wherein the nucleic acid molecule is DNA or RNA.
12. The method according to claim 9 , wherein the nucleic acid molecule is obtained from a virus.
13. The method according to claim 9 , wherein the nucleic acid molecule is single-stranded.
14. The method according to claim 9 , wherein the nucleic acid molecule is double-stranded.
15. A method of characterizing a nucleic acid molecule, the method comprising:
performing rolling circle amplification on a nucleic acid molecule of 1,000 base pairs or fewer, thereby producing a concatamer;
depositing the concatamer onto a surface such that the concatamer is elongated and fixed on the surface so that the concatamer remains accessible for enzymatic reactions;
digesting the concatamer enzymatically to produce one or more restriction digests;
imaging the restriction digests; and
constructing an optical map from the restriction digests.
16. The method according to claim 15 , wherein prior to said performing step, the method further comprises circularizing a nucleic acid molecule of 1,000 base pairs or fewer.
17. The method according to claim 15 , wherein the concatamer comprises at least five internal restriction sites.
18. The method according to claim 15 , wherein the nucleic acid molecule is DNA or RNA.
19. The method according to claim 15 , wherein the surface is derivatized glass.
20. The method according to claim 15 , wherein the nucleic acid molecule is a plasmid.
21. The method according to claim 15 , wherein the nucleic acid molecule is obtained from a virus.