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 producer cell comprising:
(a) a first construct comprising a promoter operably linked to a BIV gag, pol or gag and pol sequences;
(b) a second construct comprising a promoter operably linked to a viral envelope sequence; and
(c) a third construct comprising:
i) a DNA segment from a BIV genome,
ii) a packaging sequence to package RNA into virions,
iii) a first promoter operably linked to the DNA segment, and
iv) a transgene operably linked to a second promoter, wherein at least one of vpw, vpy and tat is deleted from said construct.
2. A producer cell comprising:
a BIV gag, poi or gag/pol coding sequence;
a viral envelope coding sequence; and
a vector construct comprising a promoter linked to 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 the promoter initiates RNA transcription of the vector construct, wherein at least one of vpw, vpy and tat is deleted from said vector construct.
3. The cell of claim 1 wherein said packaging sequence is a BIV packaging sequence.
4. The cell of claim 1 , wherein said first promoter in said third construct is an LTR promoter.
5. The cell of claim 1 wherein said first promoter in said third construct is a CMV promoter.
6. The cell of claim 1 wherein said transgene is operably linked to a CMV promoter, a PGK promoter, or an MND promoter.
7. The cell of claim 1 , wherein said third construct further comprises a rev-response element (RRE).
8. The cell of claim 1 , wherein said third construct further comprises a central polypurine tract (cPPT).
9. The cell of claim 2 , wherein said transgene is operably linked to an internal promoter.
10. The cell of claim 2 , wherein one or more nucleotides in said U3 element are mutated or deleted to diminish or eliminate U3-mediated transcription.
11. The cell of claim 2 , wherein said packaging sequence is a BIV packaging sequence.
12. The cell of claim 2 , wherein any start codons in said packaging sequence are eliminated by deletion or mutation.
13. The cell of claim 2 , wherein said U3 element further comprises a sequence that enhances polyadenylation.
14. The cell of claim 2 , wherein said vector construct further comprises a cPPT.
15. The cell of claim 14 , wherein said cPPT is a BIV cPPT.
16. The cell of claim 2 , wherein said vector construct further comprises a 3′ polypurine tract.
17. The cell of claim 2 , wherein said vector construct further comprises an RNA transport element.
18. The cell of claim 17 , wherein said RNA transport element is a lentiviral rev response element (RRE).
19. The cell of claim 18 , wherein said lentiviral RRE is a BIV RRE.
20. The cell of claim 17 , wherein said RNA transport element is a constitutive transport element (CTE).
21. The producer cell of claim 1 , wherein vpw is deleted from said construct.
22. The producer cell of claim 1 , wherein vpy is deleted from said construct.
23. The producer cell of claim 1 , wherein tat is deleted from said construct.
24. The producer cell of claim 2 , wherein vpw is deleted from said construct.
25. The producer cell of claim 2 , wherein vpy is deleted from said construct.
26. The producer cell of claim 2 , wherein tat is deleted from said construct.