Polynucleotide encoding insect ecdysone receptor
AssigneeThe Board of Trustees of the Leland Stanford University
InventorsDavid S. Hogness, Michael R. Koelle, William A. Seagraves
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Abstract
Polynucleotide sequences which encode ecdysone receptors have been isolated and expressed in host cells.
Claims45
An isolated, recombinant polynucleotide segment encoding an insect ecdysone receptor,
- wherein the ecdysone receptor binds both to an ecdysone response element and to ecdysone;
- wherein the segment comprises:
- (1) a nucleotide sequence encoding a steroid hormone receptor zinc finger DNA binding domain that binds an ecdysone response element, wherein the DNA binding domain comprises a plurality of cysteine residues, wherein the nucleotide sequence encoding the DNA binding domain hybridizes under selective hybridization conditions to nucleotides 1858 to 2055 of the DNA binding domain of Drosophila EcR of Table 2;
- and
- (2) a nucleotide sequence encoding a hormone binding domain that binds ecdysone, wherein the nucleotide sequence encoding the hormone binding domain hybridizes under selective hybridization conditions to nucleotides 2359 to 3021 of the ecdysone binding domain of Drosophila EcR of Table 2;
- wherein the selective hybridization conditions comprise a hybridization step in less than 500 mM salt and at least 37° C., and washing in 2×SSPE at 63° C.
A recombinant polynucleotide comprising a nucleic acid segment encoding a fusion protein, wherein the fusion protein comprises:
- (1) a first nucleotide sequence encoding a DNA binding domain;
- and
- (2) a second nucleotide sequence encoding a hormone binding domain that binds ecdysone, wherein the second nucleotide sequence hybridizes under selective hybridization conditions to nucleotides 2359-3021 of the ecdysone binding domain of Drosophila EcR of Table 2;
- wherein the selective hybridization conditions comprise a hybridization step in less than 500 mM salt and at least 37° C., and a washing step in 2×SSPE at 63° C., and wherein the DNA binding domain and the hormone binding domain are from different steroid receptor superfamily members.
An isolated, recombinant polynucleotide segment encoding a hormone binding domain of an insect steroid receptor, wherein the hormone binding domain binds ecdysone, wherein the polynucleotide segment encoding the hormone binding domain hybridizes to nucleotides 2359 to 3021 of the ecdysone binding domain of Drosophila EcR of Table 2 under hybridization conditions comprising a hybridization step in less than 500 mM salt and at least 37° C., and a washing step in 2×SSPE at 63° C.
An isolated, recombinant polynucleotide segment encoding a hormone binding domain of an insect ecdysone receptor, wherein the hormone binding domain exhibits saturable binding to ecdysone of an ecdysteroid, wherein the polynucleotide segment encoding the hormone binding domain hybidizes to nucleotides 2359 to 3021 of the ecdysone binding domain of Drosophila EcR of Table 2 under hybridization conditions comprising a hybridization step in less than 500 mM salt and at least 37° C., and a washing step in 2×SSPE at 63° C.
Dependencies are inferred from the claim text.