Eureka AIR delivers breakthrough ideas for toughest innovation challenges, trusted by R&D personnel around the world.

Aphelenchoides besseyi ATP synthetase and application thereof

A technology for synthesizing enzymes and nematodes, applied in the fields of application, nematicides, enzymes, etc., can solve the problems of increased mortality of second-instar larvae, weakened pathogenicity, and reduced number of host root knots, etc., to promote the expression of rice defense genes , the effect of inducing apoptosis

Active Publication Date: 2021-06-04
SOUTH CHINA AGRI UNIV
View PDF1 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Among plant nematodes, only the ATPase gene of Meloidogyne incognita has been successfully cloned. It has been reported that silencing the ATP synthase gene of Meloidogyne incognita can lead to a significant increase in the mortality of the second instar larvae of the nematode. The results of plant-mediated RNAi showed that silencing the gene resulted in weakened pathogenicity, and the number of root knots in the host was significantly reduced (Huang Y H, Mei M, Shen B M, et al. RNAi of MiASB caused high mortality of Meloidogyne incognita juveniles and inhibited the nematodediease [J].Agricultural Sciences, 2013,9(4):483-490; Huang Y H,Mei M,Mao Z C,etal.Molecular cloning and virus-induced gene silencing of MiASB in thesouthern root-knot nematode,Meloidogyne incognita[J ]. European Journal of Plant Pathology, 2014, 138:181-193)
However, there are no related reports on the ATP synthase and its function of A.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Aphelenchoides besseyi ATP synthetase and application thereof
  • Aphelenchoides besseyi ATP synthetase and application thereof
  • Aphelenchoides besseyi ATP synthetase and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1 Cloning and sequence analysis of the full-length sequence of the ATP synthase Ab-Atps gene of A.

[0038] 1. Experimental method

[0039] Using a key defense gene OsRLK3 of rice and the cDNA library of Oryza sativa, the gene was fished out through yeast two-hybrid experiment, and the partial sequence of the Ab-Atps gene of Oryza sativa was obtained by sequencing.

[0040] RACE primers were further designed through this sequence, and then using the rice A. elegans cDNA as a template, the BDSMARTTM PCR cDNA Synthesis Kit (Takara, Japan) was used to obtain the full-length gene of the rice A. elegans Ab-Atps. The specific steps were as follows:

[0041] 1. 5'RACE amplification:

[0042] (1) Primers for the first round of PCR:

[0043] Forward primer UPM: 5'-CTAATACGACTCACTCACTATAGGGC-3'

[0044] Reverse primer D2RACER1: 5'-ATCGCCAATCTGAACGATCAAGCCGCCCAT-3'

[0045] reaction system:

[0046]

[0047] Reaction conditions: 94°C for 30s, 72°C for 3min, 5 cycle...

Embodiment 2

[0070] Example 2 Determination of the expression level of the Ab-atps gene in different stages of the rice stem nematode

[0071] 1. Experimental method

[0072] 1. Use the MicroElute total RNA kit (OMEGA) to extract the RNA of 500 females, males, larvae and eggs of the test Nematode oryzae. Specific steps are as follows:

[0073] Use the MicroElute total RNA kit and follow the instructions to extract nematode RNA from small amounts. The steps are as follows:

[0074] (1) Put the nematodes (<500) into DEPC-treated 1.5mL centrifuge tubes as needed, and wash them three times with DEPC water. Place the centrifuge tube in a pre-cooled mortar, add liquid nitrogen to the mortar and immediately grind the nematodes in the tube into a fine powder with a grinding rod, and quickly add 350 μL reagent TRK and 7 μL β-mercaptoethanol.

[0075] (2) Vortex for 30s, centrifuge at 1000rpm for 1min, and pipette the supernatant into another new DEPC-treated 1.5mL centrifuge tube.

[0076] (3) ...

Embodiment 3

[0097] Example 3 In situ hybridization of Ab-atps gene

[0098] 1. Experimental method

[0099] Dilute the liquid of about 10000 mixed nematodes to 30-50 μL, add 3% paraformaldehyde solution and fix at 5 °C for 18 h, and at 22 °C for 4 h. Primers were designed according to the cDNA full-length gene of Ab-atps, and used as templates to synthesize DIG-labeled RNA probes (Roche, Germany), the specific steps were as follows:

[0100] 1. RNA probe in vitro transcription template (DNA) amplification primer

[0101] In this experiment, the RNA probe of Ab-atps was synthesized by the in vitro transcription method, and the template DNA required for in vitro transcription of the RNA probe was obtained by PCR, and the amplification primers for the template DNA were designed as follows:

[0102] A. Amplification primers for antisense strand in vitro transcription template:

[0103] IA-D2F: 5′-TAATACGACTCACTATAGGGAAATACGCGACCAGTTTGTATCAGG-3′

[0104] IA-D2R: 5′-CCGACTGTTTCAAGTTGCGAT-3′...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses Aphelenchoides besseyi ATP (adenosine triphosphate) synthetase and application thereof, the amino acid sequence of the Aphelenchoides besseyi ATP synthetase is as shown in SEQ ID NO: 2, and the nucleotide sequence of the Aphelenchoides besseyi ATP synthetase gene is shown as SEQ ID NO: 1. After RNAi treatment is adopted, the fertility of the Aphelenchoides besseyi can be inhibited, rice defense gene expression is promoted, plant immune response can be activated, and a new strategy is provided for nematode prevention and control in agricultural production. The invention provides a new target for cultivation of nematode-resistant variety plants, and provides a new way for prevention and treatment of aphelenchoides besseyi.

Description

technical field [0001] The present invention relates to the technical field of plant disease and insect pest control, in particular, to a kind of rice sharp nematode ATP synthase and application thereof. Background technique [0002] Rice is one of the most important food crops. Plant parasitic nematodes can cause economic losses of more than 16 billion US dollars in global rice every year, and the most important parasitic nematode that damages the aboveground parts of rice is rice stem-point nematode (Aphelenchoides besseyi). Oryza sativa infects rice, causing economic losses of up to 50%. The nematode is distributed in the main rice areas of my country and causes severe damage locally. [0003] ATPase gene has an important relationship with allergic necrosis induced by plant disease resistance. It has been found that the ATPase protein gene PDE1 of Magnaporthe grisea is closely related to the formation of appressoria during infection by Magnaporthe grisea; another ATPase...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N15/52C12N9/00C12N15/113A01N57/16A01P5/00
CPCC12N9/00C12N15/1137A01N57/16C12N2310/14
Inventor 谢辉丁善文王宏乐徐春玲
Owner SOUTH CHINA AGRI UNIV
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com