Claims
- 1. A shuttle vector for transforming insect cells, comprising:
a. a prokaryotic origin of replication; b. an insect promoter having homology to, and capable of functioning as, an immediate early baculovirus promoter; c. a prokaryotic promoter sequence; d. a selectable marker gene capable of conferring resistance to a bleomycin/phleomycin-type antibiotic under the transcriptional control of the insect promoter and the prokaryotic promoter sequence, in insect and prokaryotic cells respectively.
- 2. The shuttle vector of claim 1, wherein the prokaryotic promoter sequence is a cryptic promoter within the insect promoter.
- 3. The shuttle vector of claim 1, wherein the bleomycin/phleomycin-type antibiotic is Zeocin.
- 4. The shuttle vector of claim 1, further comprising an insertion site for heterologous DNA.
- 5. The shuttle vector of claim 4, wherein the insertion site for heterologous DNA is under the transcriptional control of a second insect promoter.
- 6. The shuttle vector of claim 5, further comprising a heterologous DNA sequence inserted at the insertion site and under the transcriptional control of the second insect promoter.
- 7. The shuttle vector of claim 1, wherein the insect promoter comprises an IE2B element substantially homologous to SEQ ID NO: 10.
- 8. The shuttle vector of claim 7, wherein the insect promoter comprises a GATA-IE2B element pair substantially homologous to SEQ ID NO: 9 and SEQ ID NO: 10.
- 9. The shuttle vector of claim 8, wherein the insect promoter comprises a sequence substantially homologous to SEQ ID NO: 1 from bp 351 to bp 527.
- 10. The shuttle vector of claim 9, wherein the insect promoter comprises a sequence substantially homologous to SEQ ID NO: 1.
- 11. The shuttle vector of claim 1 further comprising DNA transposable elements defining a transposon.
- 12. The shuttle vector of claim 11, wherein the selectable marker gene is within the transposon.
- 13. The shuttle vector of claim 12, further comprising an insertion site for heterologous DNA within the transposon.
- 14. The shuttle vector of claim 13, further comprising a heterologous DNA sequence inserted at the insertion site and under the transcriptional control of a second insect promoter.
- 15. The shuttle vector of claim 11, further comprising an inducible transposase gene within the transposon.
- 16. Insect cells transformed with the shuttle vector of claim 1.
- 17. Insect cells transformed with the shuttle vector of claim 11.
- 18. A method of transforming insect cells comprising:
a. inducing the insect cells to take up an insect shuttle vector comprising a selectable marker gene under the transcriptional control of an insect promoter, the selectable marker gene being capable of conferring resistance to a bleomycin/phleomycin-type antibiotic, and the insect promoter having homology to, and being capable of functioning as, an immediate early baculovirus promoter; and, b. selecting transformed cells that are resistant to the bleomycin/phleomycin-type antibiotic.
- 19. A method of increasing the copy number of a heterologous DNA sequence in a recombinant cell, comprising:
a. providing a recombinant cell having,
i. the heterologous DNA sequence positioned within a transposon defined by DNA transposable elements; ii. an unobtrusive marker gene linked to the heterologous DNA sequence within the transposon; iii. a transposase gene, the transposase being capable of mediating replicative transposition of the transposon; b. permitting expression of the transposase gene to mediate replicative transposition of the heterologous DNA sequence; and, c. monitoring the increase in copy number of the heterologous DNA sequence by monitoring the expression of the unobtrusive marker gene.
- 20. The method of claim 19 wherein the transposase gene is inducible and the step of permitting expression of the transposase gene comprises inducing expression of the transposase gene.
- 21. Recombinant insect cells transformed with the shuttle vector of claim 1, expressing a heterologous protein selected from the group consisting of melanotransferrins and biologically active derivatives thereof.
- 22. A heterologous protein produced by the cells of claim 21, selected from the group consisting of melanotransferrins and biologically active derivatives thereof.
- 23. Recombinant insect cells transformed with the shuttle vector of claim 1, expressing a heterologous protein selected from the group consisting of insect ion transport peptide hormones and biologically active derivatives thereof.
- 24. A heterologous protein produced by the cells of claim 23, selected from the group consisting of insect ion transport peptide hormones and biologically active derivatives thereof.
- 25. Insects comprising cells stably transformed with the vector of claim 1.
- 26. A recombinant cell comprising a heterologous, inducible transposase gene, wherein transposase is expressed in the cell at levels that mediate transposition only upon induction of the transposase gene.
Priority Claims (1)
Number |
Date |
Country |
Kind |
2221819 |
Jan 1998 |
CA |
|
Parent Case Info
[0001] This application claims priority to copending U.S. Provisional Application Serial No. 60/049,946 filed Mar. 27, 1997, which is incorported herein by reference.
Provisional Applications (1)
|
Number |
Date |
Country |
|
60049946 |
Mar 1997 |
US |
Continuations (1)
|
Number |
Date |
Country |
Parent |
09048911 |
Mar 1998 |
US |
Child |
09896888 |
Jun 2001 |
US |