Promoter sequences
The present invention relates to new promoter sequences and uses thereof, in particular expression cassettes, vectors, and methods of expressing genes using the new promoters.
1. An expression cassette comprising:
a polynucleotide that is:
(i) a fragment of SEQ ID NO: 1, wherein the fragment comprises nt 579 to nt 800 of the sequence as set forth in SEQ ID NO:1;
(ii) a nucleic acid sequence having at least 90% identity to (i);
(iii) the nucleic acid sequence as set forth in SEQ ID NO: 1;
(iv) a nucleic acid sequence having at least 75% identity to (iii); or
(v) the complementary sequence of (i), (ii), (iii) or (iv);
and
a heterologous coding sequence.
2. A vector comprising the expression cassette of claim 1 .
3. A method of recombinantly expressing a protein gene product, the method comprising:
expressing in a cell a gene product from an expression cassette comprising a coding sequence and a nucleic acid sequence, wherein the coding sequence is heterologous to the nucleic acid sequence and wherein said nucleic acid sequence has promoter activity and is operatively positioned for expression of the coding sequence, and wherein said nucleic acid sequence comprises
(i) the sequence as set forth in SEQ ID NO: 1 or (ii) a fragment of SEQ ID NO: 1, wherein the fragment comprises nt 450 to nt 800 or nt 579 to nt 800 of SEQ ID NO: 1.
4. A method of expressing a gene product, the method comprising
transfecting an isolated cell or cell line with an expression cassette according to claim 1 , wherein said polynucleotide has promoter activity and is operatively positioned for expression of the coding sequence, and wherein said polynucleotide comprises (i) the sequence as set forth in SEQ ID NO: 1 or (ii) a fragment of SEQ ID NO: 1, wherein the fragment comprises nt 450 to nt 800 or nt 579 to nt 800 of SEQ ID NO:1; and
expressing the gene product,
wherein, optionally, the method further comprises the step of isolating the expressed gene product.
5. The method of claim 4 , characterized in that the cell is a mammal, avian, plant or insect cell.
6. An isolated cell comprising an expression cassette according to claim 1 wherein the expression cassette is stably integrated into the genome of said cell.
7. A non-human cell comprising a vector according to claim 2 , wherein the vector is stably integrated into the genome of said cell.
8. A method of controlling expression of a gene product by an expression cassette in a cell, comprising modifying the temperature of a cell that comprises the expression cassette according to claim 1 , wherein said polynucleotide has promoter activity and is operatively positioned for expression of the coding sequence, and wherein said polynucleotide comprises (i) the sequence as set forth in SEQ ID NO: 1 or (ii) a fragment of SEQ ID NO: 1, wherein the fragment comprises nt 450 to nt 800 or nt 579 to nt 800 of SEQ ID NO: 1.
9. The expression cassette of claim 1 , wherein the fragment comprises nt 1 to nt 1334, nt 1 to nt 1294, nt 1 to nt 1206, nt 1 to nt 910, nt 450 to nt 1167, nt 450 to nt 1425, nt 115 to nt 910, nt 231 to nt 910, nt 450 to nt 910, nt 579 to nt 910, nt 1 to nt 800, nt 450 to 800, nt 115 to nt 800, nt 231 to nt 800, or nt 450 to nt 800 of SEQ ID NO:1.
10. The expression cassette of claim 1 , wherein the nucleic acid sequence of (iv) has at least 80% sequence identity to the sequence set forth in SEQ ID NO: 1.
11. The expression cassette of claim 1 , wherein the nucleic acid sequence of (iv) has at least 90% sequence identity to the sequence set forth in SEQ ID NO:1.
12. The expression cassette of claim 1 , wherein the nucleic acid sequence of (iv) has at least 95% sequence identity to the sequence set forth in SEQ ID NO:1.
13. The vector of claim 2 , wherein the fragment comprises nt 1 to nt 1334, nt 1 to nt 1294, nt 1 to nt 1206, nt 1 to nt 910, nt 450 to nt 1167, nt 450 to nt 1425, nt 115 to nt 910, nt 231 to nt 910, nt 450 to nt 910, nt 579 to nt 910, nt 1 to nt 800, nt 450 to 800, nt 115 to nt 800, nt 231 to nt 800, or nt 450 to nt 800 of SEQ ID NO:1.
14. The vector of claim 2 , wherein the nucleic acid sequence of (iv) has at least 80% sequence identity to the sequence set forth in SEQ ID NO:1.
15. The vector of claim 2 , wherein the nucleic acid sequence of (iv) has at least 90% sequence identity to the sequence set forth in SEQ ID NO: 1.
16. The vector of claim 2 , wherein the nucleic acid sequence of (iv) has at least 95% sequence identity to the sequence set forth in SEQ ID NO:1.
17. The method of claim 3 , wherein the fragment comprises nt 1 to nt 1334, nt 1 to nt 1294, nt 1 to nt 1206, nt 1 to nt 910, nt 450 to nt 1167, nt 450 to nt 1425, nt 115 to nt 910, nt 231 to nt 910, nt 450 to nt 910, nt 579 to nt 910, nt 1 to nt 800, nt 450 to 800, nt 115 to nt 800, nt 231 to nt 800, or nt 450 to nt 800 of SEQ ID NO:1.
18. The method of claim 4 , wherein the gene product is a protein.
19. The method of claim 5 , wherein the cell is a hamster cell and/or an ovarian cell.
20. The method of claim 19 , wherein the cell is a CHO cell.
21. A method of recombinantly expressing a protein gene product comprising:
expressing in a cell a gene product from an expression cassette comprising a coding sequence under the control of a nucleic acid sequence that comprises SEQ ID NO: 2, wherein the expression is induced by modifying the temperature of the cell.