Clock gene promoter
The present invention discloses a Period2 gene promoter, a construct containing the promoter and a reporter gene, a cell containing the construct, a transgenic animal harboring the construct, and a method for screening a substance which controls expression or oscillatory expression of a biological clock gene. The screening method uses the above cell, a suprachiasmatic nucleus section or peripheral tissue of the above transgenic animal or the above transgenic animal.
1. An isolated DNA which maintains a basal promoter activity and has a promoter activity transcriptionally-activated by a BMAL1/CLOCK heterodimer, which comprises the nucleotide sequence described in the following (a), (b), (c) or (d):
(a) a sequence consisting of nucleotides at positions 7,463 to 7,931 in the nucleotide sequence represented by SEQ ID NO:1,
(b) a sequence consisting of nucleotides at positions 6,417 to 7,931 in the nucleotide sequence represented by SEQ ID NO:1,
(c) a sequence consisting of nucleotides at positions 5,249 to 7,931 in the nucleotide sequence represented by SEQ ID NO:1, and
(d) a sequence consisting of nucleotides at positions 4,415 to 7,931 in the nucleotide sequence represented by SEQ ID NO:1.
2. A DNA according to claim 1 , which consists of the nucleotide sequence described in (a), (b), (c) or (d).
3. A construct which comprises the DNA according to claim 1 or 2 operably linked to a reporter gene.
4. A cell which comprises the construct according to claim 3 .
5. A method for screening a substance which controls expression of Period2 gene, comprising the steps of:
allowing the cell according to claim 4 to contact with a substance to be tested, and measuring activity of the reporter gene.
6. A transgenic rat or mouse transfected with the construct according to claim 3 , and wherein the suprachiasmatic nucleus and/or peripheral tissues of the rat or mouse exhibit the function of reporter gene expression.
7. The transgenic rat according to claim 6 .
8. A method for screening a substance which controls expression and/or oscillatory expression of Period2 gene, comprising the steps of:
allowing the cell according to claim 4 to react with a substance to be tested, and
measuring activity of the reporter gene for oscillatory expression of the Period2 gene.
9. A method for screening a substance which controls expression and/or oscillatory expression of Period2 gene, comprising the steps of:
administering a substance to be tested to the transgenic rat or mouse according to claim 6 , and
measuring activity of the reporter gene in the suprachiasmatic nucleus of the animal for oscillatory expression of the Period2 gene.
10. A method for screening a substance which controls expression and/or oscillatory expression of Period2 gene, comprising the steps of:
allowing a suprachiasmatic nucleus section or peripheral tissue of the transgenic rat or mouse according to claim 6 to react with a substance to be tested, and
measuring activity of the reporter gene for oscillatory expression of the Period2 gene.
11. The transgenic rat or mouse according to claim 6 , wherein the reporter gene is one member selected from the group consisting of a gene encoding luciferase, a gene encoding secretion type alkaline phosphatase (SEAP), a gene encoding green fluorescent protein (GFP), a gene encoding chloramphenicol acetyltransferase (CAT), a gene encoding β-glucuronidase (GUS), a gene encoding β-D-galactosidase and a gene encoding aequorin.
12. The transgenic rat according to claim 7 , wherein the reporter gene is one member selected from the group consisting of a gene encoding luciferase, a gene encoding secretion type alkaline phosphatase (SEAP), a gene encoding green fluorescent protein (GFP), a gene encoding chloramphenicol acetyltransferase (CAT), a gene encoding β-glucuronidase (GUS), a gene encoding β-D-galactosidase and a gene encoding aequorin.