Transformed cell with enhanced sensitivity to antifungal compound and use thereof
AssigneeSumitomo Chemical Company, Limited
InventorHiroki Nakajima
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Abstract
The present invention provides a transformed cell in which a polynucleotide having a nucleotide sequence encoding an amino acid sequence of an osmosensing histidine kinase having no transmembrane region is introduced in a functional form into a cell deficient in at least one hybrid-sensor kinase, a method of assaying the antifungal activity of a test substance using the transformed cell, and a method of searching an antifungal compound using the method, and the like.
Claims22
A transformed cell in which a polynucleotide having a nucleotide sequence encoding an amino acid sequence of an osmosensing histidine kinase having no transmembrane region is introduced in a functional form into a cell deficient in at least one hybrid-sensor kinase.
An osmosensing histidine kinase having no transmembrane region, wherein the kinase is derived from a plant-pathogenic filamentous fungus.
An osmosensing histidine kinase having no transmembrane region, which has an amino acid sequence selected from the group consisting of:
- (a) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which has an amino acid sequence homology of 95% or more to the amino acid sequence represented by any of SEQ ID NOs:
- 41, 55 and 68;
- (b) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which is encoded by a DNA amplified by a polymerase chain reaction using a Fusarium oxysporum-derived cDNA as a template and using an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 52 and an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 53 as primers;
- (c) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which is encoded by a DNA amplified by a polymerase chain reaction using a Mycospharella tritici-derived cDNA as a template and using an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 64 and an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 65 as primers;
- (d) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which is encoded by a DNA amplified by a polymerase chain reaction using a Thanapethorus cucumeris-derived cDNA as a template and using an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 85 and an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 86 as primers;
- (e) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which is derived from Phytophthora infestans and has the amino acid sequence represented by SEQ ID NO:
- 90;
- (f) the amino acid sequence represented by SEQ ID NO:
- 41;
- (g) the amino acid sequence represented by SEQ ID NO:
- 55;
- and
- (h) the amino acid sequence represented by SEQ ID NO:
- 68.
An osmosensing histidine kinase having no transmembrane region, which has an amino acid sequence represented by SEQ ID NO:
- 41, SEQ ID NO:
- 55 or SEQ ID NO:
- 68.
A polynucleotide having a nucleotide sequence encoding an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, wherein the kinase is derived from a plant-pathogenic filamentous fungus.
A polynucleotide having a nucleotide sequence encoding an amino acid sequence selected from the group consisting of:
- (a) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which has an amino acid sequence homology of 95% or more to the amino acid sequence represented by any of SEQ ID NOs:
- 41, 55 and 68;
- (b) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which is encoded by a DNA amplified by a polymerase chain reaction using a Fusarium oxysporum-derived cDNA as a template and using an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 52 and an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 53 as primers;
- (c) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which is encoded by a DNA amplified by a polymerase chain reaction using a Mycospharella tritici-derived cDNA as a template and using an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 64 and an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 65 as primers;
- (d) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which is encoded by a DNA amplified by a polymerase chain reaction using a Thanapethorus cucumeris-derived cDNA as a template and using an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 85 and an oligonucleotide having the nucleotide sequence represented by SEQ ID NO:
- 86 as primers;
- (e) an amino acid sequence of an osmosensing histidine kinase having no transmembrane region, which is derived from Phytophthora infestans and has the amino acid sequence represented by SEQ ID NO:
- 90;
- (f) the amino acid sequence represented by SEQ ID NO:
- 41;
- (g) the amino acid sequence represented by SEQ ID NO:
- 55;
- and
- (h) the amino acid sequence represented by SEQ ID NO:
- 68.
A polynucleotide having a nucleotide sequence represented by SEQ ID NO:
- 42, SEQ ID NO:
- 56 or SEQ ID NO:69.
A method of obtaining a polynucleotide having a nucleotide sequence encoding an amino acid sequence of osmosensing histidine kinase which is derived from a plant-pathogenic filamentous fungus and has no transmembrane region, which comprises a step of amplifying a desired polynucleotide by Polymerase Chain Reaction using an oligonucleotide having a nucleotide sequence represented by any of SEQ ID NOs:
- 30 to 40, 52, 53, 64, 65, 85 and 86 as primers, and a step of recovering the amplified desired polynucleotide.
An oligonucleotide which comprises a nucleotide sequence represented by any of SEQ ID NOs:
- 30 to 40, 52, 53, 64, 65, 85 and 86.
Dependencies are inferred from the claim text.