Lepidoptera pest ground substance metalloprotein gene mmp and application thereof

A matrix metal, lepidopteran technology, applied in the field of regulating the allergic response of lepidopteran pests, can solve the problem of little research

Inactive Publication Date: 2008-07-02
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are few studies on the gene and its products in Lepidoptera pests such as tea geometrids

Method used

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  • Lepidoptera pest ground substance metalloprotein gene mmp and application thereof

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Experimental program
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Effect test

Embodiment 1

[0028] Take 2-3 tea geometrid larvae in the feeding period of the 3rd instar and the dormancy period at the end of the 3rd instar respectively. Total RNA was extracted with TRIZOL (Invitrogen Company) reagent, and mRNA was isolated with UNIQ-10 column mRNA extraction and purification kit (Shanghai Sangon Bioengineering Co., Ltd.). 1 μg of mRNA was used for cDNA synthesis, and double-stranded cDNA was synthesized using TakaRa MMLV RTase cDNASynthesis Kit (Takara Bioengineering [Dalian] Co., Ltd.). The double-stranded cDNA was purified by isopropanol precipitation and digested with TakaRa Afl II / Nsp I (Takara Bioengineering [Dalian] Co., Ltd.). The digested fragment was ligated with two previously designed linkers, SEQ ID No: 3 and SEQ ID No: 4, with T4 DNA ligase (Shanghai Sangon Bioengineering Co., Ltd.). Use the ligated cDNA fragment as a template, and perform PCR pre-amplification with pre-amplification primers SEQ ID No: 5 and SEQ ID No: 6. After diluting the pre-amplifica...

Embodiment 2

[0031]Tea geometrid mmp SEQ ID No: 1 was used as the material, and TaKaRa siRNA Cocktail Kit (si-RNAase IIITM) (Treasure Bioengineering [Dalian] Co., Ltd.) was used to prepare the mmp that can undergo double-strand interference with the endogenous mmp of Lepidoptera pests -siRNAs. Tea geometrids that had just entered the prepupal stage were divided into two groups, one group was treated with mmp-siRNAs, and the other group was control. After dissolving the prepared mmp-siRNAs in water, inject the mmp-siRNAs in the thorax and abdomen of the anterior pupae, and the injection dose is 150ng (volume is 2 μ L) / head, that is, each head injects 150ng of mmp-siRNAs; control. After 48 hours of injection treatment, two animals were selected from the treatment group and the control group respectively, and the total RNA was extracted with TRIZOL (Invitrogen). Using SEQ ID No: 10 and SEQ ID No: 11 as primers, RT-PCR was performed with the MMLV One-Step RT-PCR Amplification Kit. For specif...

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Abstract

The invention discloses a matrix metalloproteinase gene (mmp) derived from a Lepidoptera pest, whic has the nucleotide sequence shown in SEQ ID NO: 1. The invention also discloses a protein encoded by the gene mmp, which has an amoni acid sequence shown in SEQ ID NO: 2. The invention further discloses the use of the gene mmp in regulation of allergic reaction of Lepidoptera pests.

Description

technical field [0001] The invention belongs to the field of insect biotechnology, and in particular relates to the separation of the tea geometrid matrix metalloproteinase gene and the use of the gene to regulate the allergic reaction of the lepidopteran pests. Background technique [0002] Insects of Lepidoptera, such as tea geometrid, are one of the important pests of tea trees. Effective control of these pests is the fundamental guarantee for high yield and high quality of tea. However, with the increase in the frequency and dosage of existing pesticides, the resistance of Lepidoptera pests gradually increases, resulting in a decline in the control effect. Therefore, it is necessary to develop some new pesticides with strong pertinence, clear action sites and good control effects. [0003] Matrix metalloproteinase (mmp) is the most important proteolytic system for biodegrading extracellular matrix components and plays an important role in tissue remodeling. MMPs are a ...

Claims

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Application Information

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IPC IPC(8): C12N15/57C12N9/64C12Q1/68
Inventor 陆建良林晨梁月荣郑新强杜颖颖
Owner ZHEJIANG UNIV
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