Aptamer constructs
View Patent ↗Provided are RNA oligonucleotides having an aptamer flanked by two self-cleaving ribozymes. Vectors encoding the oligonucleotides, and cells transfected with these vectors are also provided. Additionally, methods of inhibiting replication of a virus in a cell, methods of treating an organism with an aptamer, methods of determining whether a test aptamer is effective in inhibiting the function of a target of the aptamer in a cell, and methods of expressing an RNA aptamer in a cell are provided.
1. An RNA oligonucleotide comprising an aptamer flanked by two self-cleaving ribozymes, wherein the aptamer comprises the sequence set forth in SEQ ID NO:2, SEQ ID NO:3 or SEQ ID NO:6.
2. An RNA oligonucleotide comprising an aptamer flanked by two self-cleaving ribozymes, wherein the self-cleaving ribozymes comprise SEQ ID NO:9 or SEQ ID NO:10.
3. The RNA oligonucleotide of claim 2 , further comprising a third ribozyme between the two self-cleaving ribozymes and adjacent to the aptamer, wherein the third ribozyme is not self-cleaving.
4. A vector encoding the RNA oligonucleotide of claim 2 , wherein the vector further comprises a promoter directing transcription of the RNA oligonucleotide.
5. The vector of claim 4 , wherein the vector is a retroviral vector.
6. The vector of claim 4 , wherein the vector is a pBabe MuLV retroviral vector and the promoter is a tRNA val promoter.
7. The vector of claim 4 , further comprising a viral long terminal repeat (LTR) driving expression of the vector, wherein the aptamer is in reverse orientation to the LTR, and wherein the vector is an MMP-eGFP vector.
8. A cell transfected with the vector of claim 4 .
9. The cell of claim 8 , wherein the cell is a mammalian cell.
10. The cell of claim 8 , wherein the cell is a human cell.
11. The cell of claim 8 , wherein the aptamer is capable of inhibiting replication of a virus in the cell.
12. The cell of claim 11 , wherein the cell is infected with the virus.
13. The cell of claim 11 , wherein the virus is a retrovirus.
14. The cell of claim 11 , wherein the virus is HIV-1.
15. A cell expressing an aptamer, wherein the aptamer does not comprise flanking sequences that interfere with the aptamer function, wherein the aptamer binds to a component of a retrovirus, and wherein the aptamer comprises the sequence set forth in SEQ ID NO:2, SEQ ID NO:3 or SEQ ID NO:6 and/or the aptamer is cleaved from an RNA oligonucleotide comprising an RNA sequence flanked by two self-cleaving ribozymes that comprise SEQ ID NO:9 or SEQ ID NO:10.
16. The cell of claim 15 , wherein the cell is infected with HIV-1 virus.
17. The cell of claim 15 , wherein the aptamer inhibits replication of HIV-1 virus when the aptamer and the virus are present in the cell.
18. The RNA oligonucleotide of claim 2 , wherein the ribozymes recognize a GUC cleavage motif adjacent to the aptamer.
19. The cell of claim 17 , wherein the ribozymes recognize a GUC cleavage motif adjacent to the aptamer.
20. The RNA oligonucleotide of claim 1 , further comprising a third ribozyme between the two self-cleaving ribozymes and adjacent to the aptamer, wherein the third ribozyme is not self-cleaving.
21. A vector encoding the RNA oligonucleotide of claim 1 , wherein the vector further comprises a promoter directing transcription of the RNA oligonucleotide.
22. The vector of claim 21 , wherein the vector is a retroviral vector.
23. The vector of claim 21 , wherein the vector is a pBabe MuLV retroviral vector and the promoter is a tRNA val promoter.
24. The vector of claim 21 , further comprising a viral long terminal repeat (LTR) driving expression of the vector, wherein the aptamer is in reverse orientation to the LTR, and wherein the vector is an MMP-eGFP vector.
25. A cell transfected with the vector of claim 21 .
26. The cell of claim 25 , wherein the aptamer is capable of inhibiting replication of a virus in the cell.
27. The cell of claim 25 , wherein the cell is infected with the virus.
28. The cell of claim 27 , wherein the virus is a retrovirus.
29. The cell of claim 28 , wherein the virus is HIV-1.
30. The cell of claim 25 , wherein the cell is a mammalian cell.
31. The cell of claim 25 , wherein the cell is a human cell.
32. The RNA oligonucleotide of claim 1 , wherein the ribozymes recognize a GUC cleavage motif adjacent to the aptamer.