Bovine immunodeficiency virus (BIV) based vectors
This invention pertains to BIV constructs encompassing BIV combination vectors, BIV vectors and BIV packaging vectors and particularly the invention pertains to a three vector system comprising: a) a BIV vector construct including a DNA segment from a BIV genome, a packaging sequence to package RNA into virions; a promoter operably linked to the DNA segment; and a transgene operably linked to a second promoter; b) a BIV packaging vector construct comprising a BIV DNA sequence fragment comprising at least a gag gene or pol gene of BIV; a promoter operably linked to the BIV DNA fragment; and a polyadenylation sequence located downstream of the BIV DNA fragment; and c) an expression vector construct comprising a gene encoding a viral surface protein. Also provided is a method for transferring a gene of interest into a mammalian cell.
1. A method of transferring a transgene to a mammalian cell comprising administering a vector particle comprising an RNA, comprising:
a first BIV R region;
a BIV U5 element linked to the first BIV R region;
a packaging sequence;
a transgene; and
a BIV U3 element linked to a second BIV R region, wherein coding sequences of at least one of vpw, vpy and tat is deleted from said RNA.
2. The method of claim 1 , wherein said transgene is operably linked to an internal promoter.
3. The method of claim 1 , wherein one or more nucleotide sequences in said U3 element are mutated or deleted to diminish or eliminate U3-mediated transcription.
4. The method of claim 1 , wherein said U3 element further comprises a sequence that enhances polyadenylation.
5. The method of claim 1 , wherein said packaging sequence is a BTV packaging sequence.
6. The method of claim 1 , wherein any start codons in said packaging sequence are rendered non-functional.
7. The method of 1 , wherein said RNA further comprises a BIV cPPT.
8. The method of claim 1 , wherein said RNA further comprises an RNA transport element.
9. The method of claim 8 , wherein said RNA transport element is a lentiviral rev response element (RRE).
10. The method of claim 9 , wherein said lentiviral RRE is a BIV RRE.
11. The method of claim 8 , wherein said RNA transport element is a constitutive transport element (CTE).
12. A method of transferring a transgene to a mammalian cell comprising administering a BIV vector particle comprising:
a) an RNA segment from a BIV genome,
b) a packaging sequence to package RNA into virions, and
c) a transgene operably linked to a promoter, wherein at least one of vpw, vpy and tat is deleted from said construct.
13. The method of claim 12 , wherein said packaging sequence is a BIV packaging sequence.
14. The method of claim 12 , wherein said promoter is a mammalian promoter.
15. The method of claim 12 wherein said promoter is a CMV promoter, a PGK promoter, or an MND promoter.
16. The method of claim 12 , said RNA segment further comprising a rev-response element.
17. The method of claim 12 , said RNA segment further comprising a central polypurine tract.