Compositions and methods of nucleic acid-targeting nucleic acids
This disclosure provides for compositions and methods for the use of nucleic acid-targeting nucleic acids and complexes thereof. Genome engineering can refer to altering the genome by deleting, inserting, mutating, or substituting specific nucleic acid sequences. The altering can be gene or location specific. Genome engineering can use nucleases to cut a nucleic acid thereby generating a site for the alteration. Engineering of non-genomic nucleic acid is also contemplated.
1. A method for detecting if two complexes are in proximity to one another comprising:
contacting a first target nucleic acid with a first complex, wherein said first complex comprises a first site-directed polypeptide, a first modified nucleic acid-targeting nucleic acid comprising a first nucleic acid sequence complementary to the first target nucleic acid, and a first effector protein, wherein said first effector protein is adapted to bind to said first modified nucleic acid-targeting nucleic acid, and wherein said first effector protein comprises a first non-native sequence that comprises a first portion of a split system;
contacting a second target nucleic acid with a second complex, wherein said second complex comprises a second site-directed polypeptide, a second modified nucleic acid-targeting nucleic acid comprising a second nucleic acid sequence complementary to the second target nucleic acid, and a second effector protein, wherein said second effector protein is adapted to bind to said second modified nucleic acid-targeting nucleic acid, and wherein said second effector protein comprises a second non-native sequence that comprises a second portion of a split system; and
detecting an interaction between said first portion and said second portion, wherein said detecting indicates said first complex and said second complex are in proximity to one another.
2. The method of claim 1 , wherein said split system comprises two or more protein fragments that individually are not active but, when formed into a complex, result in an active protein complex.
3. The method of claim 1 , wherein said detecting comprises determining the occurrence of a genetic mobility event.
4. The method of claim 3 , wherein said genetic mobility event comprises a translocation.
5. The method of claim 3 , wherein prior to said genetic mobility event said two portions of said split system do not interact.
6. The method of claim 3 , wherein after said genetic mobility event said two portions of said split system do interact.
7. The method of claim 3 , wherein said genetic mobility event is a translocation between a BCR and an Abl gene.
8. The method of claim 1 , wherein said split system is selected from the group consisting of a split GFP system, a split ubiquitin system, a split transcription factor system, a split affinity tag system, and any combination thereof.
9. The method of claim 1 , wherein said detecting indicates a genotype.
10. The method of claim 1 , wherein said first target nucleic acid and said second target nucleic acid are DNA.
11. The method of claim 1 , wherein said first modified nucleic acid-targeting nucleic acid and said second modified nucleic acid-targeting nucleic acid comprise RNA.
12. The method of claim 1 , wherein said first site-directed polypeptide and said second site-directed polypeptide comprise Cas9.