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Agent that modulates physiological condition of pests, involved in insect voltage-gated potassium channel activity

A technology of potassium channel and insecticidal activity, applied in the field of reagents involved in insect voltage-gated potassium channel activity to regulate the physiological status of pests, and can solve problems such as neuromuscular defects, gain-of-function mutations, movement and oviposition defects, etc.

Inactive Publication Date: 2011-10-19
SUMITOMO CHEM CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Loss-of-function mutants of the seizure ortholog unc-103 of Caenorhabditis elegans do not display overt defects, but gain-of-function (channel activation) mutations result in severe motility and egg-laying defects (WormBase, www. wormbase.org / )
These data suggest that hyperactivation of seizure-like potassium channels also leads to severe neuromuscular deficits

Method used

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  • Agent that modulates physiological condition of pests, involved in insect voltage-gated potassium channel activity
  • Agent that modulates physiological condition of pests, involved in insect voltage-gated potassium channel activity
  • Agent that modulates physiological condition of pests, involved in insect voltage-gated potassium channel activity

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0421] Example 1 (total RNA is extracted from cotton aphid and German cockroach)

[0422] (1) Extract total RNA from cotton aphid

[0423] Adults and larvae of a colony of cotton aphids (Aphis gossypii) cultivated on leaves of potted cucumbers were scraped from the leaf surfaces with a small brush, and 630 mg of the resulting colonies were pressed into powder with a mortar and pestle in liquid nitrogen. From the obtained freeze-crushed powder, RNA was isolated using RNA extraction reagent ISOGEN (manufactured by Nippon Gene) according to the following. After adding 10 ml of ISOGEN to the frozen crushed powder in the mortar, the crushed powder was kept on ice for 10 minutes for grinding. After milling, the fluid sample was transferred to a 15 ml test tube with a pipette, and 2 ml of chloroform (manufactured by Wako Pure Chemical Industries, Ltd.) was added thereto. The mixture was immediately shaken vigorously for 15 seconds, then allowed to stand at room temperature for 3 mi...

Embodiment 2

[0426] Embodiment 2 (isolation cotton aphid seizure gene)

[0427] Using total RNA from cotton aphids, random primers for RT-PCR (Invitrogen) and Superscript III (Invitrogen), first-strand cDNA was prepared according to the manufacturer's protocol of Superscript III.

[0428] By using an oligonucleotide comprising the nucleotide sequence of SEQ ID NO: 11 and an oligonucleotide comprising the nucleotide sequence of SEQ ID NO: 12, which are primers specific to the gene, and extending the long template PCR The system (manufactured by Roche) performs PCR according to the manufacturer's method to amplify the full-length cDNA of cotton aphid seizure. The cDNA described in Example 1 was used as a template. The PCR conditions used were those for descending PCR, as follows: initial denaturation at 94° C. for 2 minutes; then 10 descending-PCR cycles, one cycle being 94° C. for 30 seconds, and 70° C. for 30 seconds (each cycle decreased by 1 °C), and 68 °C for 4 minutes; followed by 25...

Embodiment 3

[0429] Embodiment 3 (separate german cockroach seizure gene)

[0430] In order to isolate homologues of the seizure gene from other insect species such as the German cockroach (Blatella germanica), the Codehop program (Blocks Protein Analysis Server (Blocks Protein Analysis Server) run by the Fred Hutchinson Cancer Research Center) was applied. website http: / / blocks.fhcrc.org / blocks / codehop.html), and with reference to the amino acid sequence of the aforementioned cotton aphid seizure and previously known amino acid sequences from humans such as (NCBI accession numbers Q12809 and Q9H252 ), Caenorhabditis elegans (NP_49782), Drosophila melanogaster (AAM68296), and Anopheles gambiae (XP_308166) homologue amino acid sequences, and degenerate primers were designed.

[0431] A partial sequence of a homologue of the seizure gene of a selected insect species is amplified by a series of PCRs using the first-strand cDNA derived from the insect species as a template. Here, the first-st...

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Abstract

The present invention provides an agent that modulates physiological condition of pests, wherein the agent has an ability to modulate the activity of an insect voltage-gated potassium channel; a method for assaying pesticidal activity of a test substance, which comprises measuring the activity of a voltage-gated potassium channel in a reaction system in which the voltage-gated potassium channel contacts with a test substance; and so on.

Description

technical field [0001] The invention relates to a reagent for regulating the physiological state of pests, participating in the activity of insect voltage-gated potassium channels, and the like. Background technique [0002] Voltage-gated potassium channels are membrane proteins that respond to membrane potential to selectively permeate potassium ions and are ubiquitous in animals, plants, and microorganisms, and include a variety of molecular species. According to the structural and physiological functional characteristics, the voltage-gated potassium channels are classified into many families, namely Shaker type, EAG (ether-a-go-go) type, KvLQT, etc. (Gutman et al., Pharmacol. Reviews (Pharmacological Reviews) , 55(4):583-586, 2003). [0003] Seizure is the insect homologue of human ERG (hERG) and a member of the EAG family of voltage-gated potassium channels. The EAG family consists of three subfamilies EAG (ether-a-go-go), ERG (EAG-related genes) and ELK (EAG-like gene...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/435G01N33/94
CPCC07K14/43563G01N33/5085G01N33/6872G01N2500/10
Inventor 大槻纯子马克·洛吉泰特斯·卡莱塔温迪·马德雷恩
Owner SUMITOMO CHEM CO LTD
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