Isolation of novel AAV's and uses thereof
The invention in some aspects relates to isolated nucleic acids, compositions, and kits useful for identifying adeno-associated viruses in cells. In some aspects, the invention provides kits and methods for producing somatic transgenic animal models using recombinant AAV (rAAV) to an animal having at least one transgene that expresses a small interfering nucleic acid or at least one binding site for a miRNA.
1. A kit for producing a rAAV that generates a somatic transgenic animal that expresses a transgene in a target tissue, comprising:
at least one container housing a recombinant AAV vector, wherein the recombinant AAV vector comprises a transgene that expresses an RNA transcript that comprises at least one binding site for a miRNA that is expressed in cells of an off-target tissue of the animal, wherein, when the miRNA and the RNA transcript are present together in cells of the off-target tissue, the miRNA hybridizes to the at least one binding site of the RNA transcript and the miRNA inhibits activity of the RNA transcript wherein the miRNA is miR-122 and the off-target tissue is liver;
at least one container housing a rAAV packaging component, wherein the packing component comprises a host cell comprising at least one cap gene encoding a capsid protein other than AAV9 serotype capsid protein, and;
instructions for constructing and packaging the rAAV, wherein the rAAV transduces cells of the target tissue.
2. The kit of claim 1 , wherein the host cell further comprises at least one rep gene.
3. The kit of claim 1 , wherein the AAV capsid protein serotype is selected from: AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV10, AAV11, AAV12, and variants thereof.
4. The kit of claim 1 , wherein the RNA transcript is an exogenous miRNA.
5. The kit of claim 1 , wherein the transcript is an mRNA transcript that encodes a protein.
6. The kit of claim 1 , wherein the transgene comprises a tissue specific promoter or inducible promoter operatively linked to a coding region from which the RNA transcript is expressed.
7. The kit of claim 1 , wherein the rAAV has an Inverted Terminal Repeat (ITR) sequence from an AAV serotype selected from: AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, and AAV12, and variants thereof.
8. The kit of claim 1 , wherein the product of the transgene causes a pathological state in the animal.
9. The kit of claim 1 , wherein product of the transgene is a reporter molecule.
10. A recombinant adeno-associated virus (rAAV) comprising:
an AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV10, AAV11, or AAV12 capsid in which is packaged a transgene that encodes an RNA transcript that comprises at least one binding site for miR-122 that inhibits expression of the RNA transcript in liver.
11. The rAAV of claim 10 , wherein the transgene comprises up to three binding sites for miR-122.
12. The rAAV of claim 10 , wherein the RNA transcript is an exogenous miRNA.
13. The rAAV of claim 10 , wherein the transcript is an mRNA transcript that encodes a protein.
14. The rAAV of claim 10 , wherein the rAAV has an Inverted Terminal Repeat (ITR) sequence from an AAV serotype selected from: AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV10, AAV11, and AAV12, and variants thereof.
15. The rAAV of claim 10 , wherein the transgene comprises a tissue-specific promoter operatively linked to a coding region from which the RNA transcript is expressed.