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Oedaleus asiaticus extracellular regulated protein kinase (ERK) as well as encoding gene and application thereof

A kind of car locust, protease technology, applied in application, genetic engineering, plant genetic improvement and other directions, can solve problems such as harming the environment, and achieve the effects of weight loss and overall performance reduction.

Active Publication Date: 2021-01-15
宁夏中微泰克生物技术有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The purpose of this application is to provide a kind of Asiatic locust exoproteinase ERK and its coding gene and application, in order to solve the problem of harmful environment caused by chemical pesticide control insect pests

Method used

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  • Oedaleus asiaticus extracellular regulated protein kinase (ERK) as well as encoding gene and application thereof
  • Oedaleus asiaticus extracellular regulated protein kinase (ERK) as well as encoding gene and application thereof
  • Oedaleus asiaticus extracellular regulated protein kinase (ERK) as well as encoding gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] Embodiment 1, the acquisition of the extracellular protease ERK and its coding gene of P. asiaticus

[0091] 1. Extraction of total RNA from P. asiaticus

[0092] use The RNA isolation reagent was used to extract the RNA from the tissue samples of P. trocarpus. Specific steps are as follows:

[0093] 1) Put a 2mL homogenizer in an oven, sterilize at 160°C for 3 hours, and cool to room temperature for later use.

[0094] 2) Put the homogenizer on ice, add 1mL RNA isolation reagent and 100-200 mg of P. asiaticus tissue, ground.

[0095] 3) Transfer the homogenate to a 1.5mL centrifuge tube and let stand at room temperature for 5min. Centrifuge at 13000r for 5min at 4°C.

[0096] 4) Transfer the supernatant to a clean 1.5mL centrifuge tube, add 200μL chloroform, and vortex for 15s. Place at room temperature for 5 minutes. Centrifuge at 13000r for 10min at 4°C.

[0097] 5) Transfer 400 μL of supernatant to a new 1.5 mL centrifuge tube, add 200 μL of chloroform,...

Embodiment 2

[0130] Embodiment 2, the dsRNA of the extracellular protease gene ERK and its application in preventing and controlling pests

[0131] 1. Synthesis of dsRNA

[0132] dsRNA was synthesized using T7 RiboMAX™ Express RNAi System kit. Specific steps are as follows:

[0133] 1) Synthesis of dsRNA primers

[0134] Design primers based on the cloned gene fragments, amplify the target fragment to about 500 bp, and introduce T7 promoter at the 5' end of the primers. The primer sequences are as follows:

[0135] erk-2F: 5'-TAATACGACTCACTATAGGAACTGAGCAACGACCACA-3';

[0136] erk-2R: 5'-TAATACGACTCACTATAGGGAGCGTCTTCAACAACTATCC-3'.

[0137] 2) Preparation of DNA template

[0138] The bacterial liquid plasmid was extracted with a kit, and the plasmid containing the gene fragment (recombinant vector in Example 1) was used as a template, and ERK-2F and ERK-2R were used for PCR amplification to obtain the target fragment containing the T7 promoter sequence.

[0139] The PCR reaction syst...

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Abstract

The invention discloses an oedaleus asiaticus extracellular regulated protein kinase (ERK) as well as an encoding gene and application thereof. The method comprises the following steps: cloning the oedaleus asiaticus ERK gene from oedaleus asiaticus, designing primers for the oedaleus asiaticus ERK gene, synthesizing dsRNA for interfering with the oedaleus asiaticus ERK gene, and introducing the dsRNA into oedaleus asiaticus by an injection method to perform RNAi on the oedaleus asiaticus ERK gene. Results show that after dsRNA is injected into the oedaleus asiaticus, the survival rate, the growth rate, the weight and the overall expressive force are all remarkably reduced, and it is indicated that the oedaleus asiaticus ERK gene plays an important role in the insect growth and developmentprocess. A novel method is provided for molecular regulation of pests, and a theoretical basis is provided for deep understanding of the insect growth and development mechanism and creation of new biopesticide preparations.

Description

technical field [0001] The application relates to the technical field of biotechnology, in particular to an extracellular protease ERK of the Asian carcus spp. and its coding gene and application. Background technique [0002] In recent years, the degree of harm of grassland pests on grasslands has shown an obvious upward trend, and the area of ​​endangered grasslands has increased year by year, seriously threatening agricultural and animal husbandry production, and also laying hidden dangers for the occurrence of sandstorms. Traditional pest control is mostly based on emergency control and chemical control, and the control methods rely too much on chemical pesticides. This not only endangers the environment, but also issues such as pesticide residues are also key topics that people are increasingly concerned about. This shows that the current stage of crop pest control There are disadvantages of relative lag. Therefore, pest control is a long process, which needs to be car...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/64C12N15/57C12N15/113C12N15/89A01N57/16A01P7/04
CPCC12N9/6405C12N15/1137C12N15/89A01N57/16C12N2310/14
Inventor 涂雄兵张泽华李霜徐超民
Owner 宁夏中微泰克生物技术有限责任公司
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