Expression vectors comprising the mCMV IE2 promoter
The invention relates to an expression vector comprising the promoter of the mCMV-IE2 gene, or a functional expression promoting fragment thereof, and/or an enhancer of the mCMV-IE2 gene, or a functional expression enhancing fragment thereof, wherein the expression vector does not contain any complete gene of the mCMV.
1. An expression vector comprising a mCMV-IE2 promoter fragment consisting of nucleotides 1 to 39 of SEQ ID NO: 1 directly linked to a DNA sequence encoding a heterologous polypeptide.
2. The vector according to claim 1 , wherein said vector further comprises a mCMV-IE2 enhancer comprising SEQ ID NO: 9.
3. The vector according to claim 2 , wherein said mCMV-IE2 enhancer comprises SEQ ID NO: 10.
4. The vector according to claim 1 , further comprising a second promoter different from said mCMV-IE2 promoter.
5. The vector according to claim 4 , wherein the second promoter is the mCMV-IE1 promoter.
6. The vector according to claim 4 , wherein the mCMV-IE2 promoter fragment and the second promoter are bi-directionally arranged.
7. The vector according to claim 4 , further comprising one or more regulatory elements selected from a 5′UTR, an intron, a 3′UTR, a mRNA 3′ end processing sequence, a polyadenylation site, or an internal ribosome entry sequence (IRES).
8. The vector according to claim 7 , wherein the IRES is operably linked to a DNA sequence encoding at least one polycistronic mRNA.
9. The vector according to claim 7 , further comprising one or more DNA element selected from an insulator, a boundary element, a locus control region (LCR), a matrix attachment region (MARs), an element for recombination or an element for cassette exchange.
10. A host cell transfected with the vector according to claim 1 .
11. The host cell according to claim 10 , wherein the host cell is a CHO cell.
12. A process for the production of a polypeptide comprising the step of transfecting a host cell according to claim 10 and culturing said host cell under conditions allowing expression of the polypeptide.
13. The process according to claim 12 , wherein the transfection is a stable transfection.
14. The process according to claim 13 , further comprising the step of isolating the polypeptide from the host cell or cell culture supernatant.
15. The process according to claim 12 , further comprising the step of isolating the polypeptide from the host cell or cell culture supernatant.
16. The vector according to claim 1 , wherein said expression vector further comprises a second promoter directly linked to a DNA sequence encoding a second polypeptide.
17. A host cell comprising an expression vector according to claim 16 .
18. The vector according to claim 16 , wherein the second promoter is different from said mCMV-IE2 promoter.
19. The vector according to claim 16 , wherein the second promoter is the mCMV-IE1 promoter.
20. The vector according to claim 16 , wherein the heterologous polypeptide or the second polypeptide is a marker protein.
21. The vector according to claim 20 , wherein the marker protein is selected from an adenosine deaminase (ADA), an aminoglycoside phosphotransferase (neo), a dihydrofolate reductase (DHFR), a hygromycin-B-phosphotransferase (HPH), a thymidine kinase (tk), a xanthine-guanine phosphoribosyltransferase (gpt), a multiple drug resistance gene (MDR), an ornithine decarboxylase (ODC), carbamyl- P synthetase/aspartate transcarbamylase/dihydro-orotase, a puromycin acetyltransferase (PAC), a galactokinase, a human folate receptor, or a reduced folate carrier.
22. The vector according to claim 16 , wherein the heterologous polypeptide or the second polypeptide is a reporter protein.
23. The vector according to claim 22 , wherein the reporter protein is selected from a luciferase, a green fluorescent protein, an alkaline phosphatase, and a horseradish peroxidase or combinations thereof.
24. The vector according to claim 16 , wherein the heterologous polypeptide and the second polypeptide are the same.
25. The vector according to claim 16 , wherein the heterologous polypeptide and the second polypeptide are different.
26. The vector according to claim 25 , wherein the heterologous polypeptide is a first subunit of a dimeric or a multimeric protein and the second polypeptide is a second subunit of a dimeric or a multimeric protein.
27. The vector according to claim 26 , wherein the first and the second subunit comprise the alpha and the beta chain of a hormone selected from a human FSH, a human LH, a human TSH or a human CG.
28. The vector according to claim 26 , wherein one subunit is a heavy chain and the other subunit is a light chain of an immunoglobulin.