Insect chitin synthase 1B gene segment, dsRNA and application thereof

A technology of chitin synthase and gene fragments, applied in the direction of DNA/RNA fragments, applications, recombinant DNA technology, etc., can solve the problems of non-target biological impact, long killing time, environmental pollution, etc.

Inactive Publication Date: 2010-09-15
SHANXI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0002] The damage caused by agricultural pests is an important factor restricting agricultural production in my country. Long-term application of chemical insecticides leads to a series of problems: 1) Insects become resistant to insecticides, the dosage of pesti...

Method used

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  • Insect chitin synthase 1B gene segment, dsRNA and application thereof
  • Insect chitin synthase 1B gene segment, dsRNA and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0013] Example 1: Acquisition of Chitin Synthetase 1B Gene Fragment and dsRNA of East Asian Migratory Locust

[0014] 1) Acquisition of chitin synthase 1B gene fragment of Migratory locust

[0015] According to the nucleotide sequence of migratory locust chitin synthase 1B with the accession number GU067731 in the NCBI database, specific primers were designed using primer premier5.0 software. The sequence of the upstream primer is SEQ ID NO: 2, and the sequence of the downstream primer is SEQ ID NO: 2. ID NO: 3, all primers were synthesized by Shanghai Yingwei Jieji Biological Co., Ltd. Select the 5th instar nymphs of migratory locust with equal size, half male and half male, in groups of four, and freeze them in liquid nitrogen for RNA extraction. For the specific operation steps of RNA extraction, refer to the TaKaRa Trizol kit. M-MLV reverse transcriptase reverse-transcribes the extracted RNA into the first-strand cDNA, and uses this as a template to amplify the chitin syn...

Embodiment 2

[0018] Example 2: The dsRNA synthesized by the chitin synthase 1B gene fragment killed the East Asian migratory locust experiment

[0019] 1. Injection of dsRNA synthesized by chitin synthase 1B gene fragment of migratory locust

[0020] The nymphs of the 2nd instar migratory locust with uniform size and health status on the fourth day were selected for injection of the above-mentioned synthetic dsRNA. A 25 μl micro-syringe is used for injection. Do not use too much force when injecting. Follow the direction of blood flow. The junction of the 2nd to 3rd abdominal segment of the flank is used as the injection point, avoiding the valve. The amount of dsRNA injected with the chitin synthase 1B gene fragment was 3 μg, and a control group of dsGFP (3 μg) was set up, with 15 worms in each group, 3 biological replicates, a total of 45 worms. After the injection, the worms were reared in a 1L beaker in an artificial climate box (light:dark time=14h:10h, temperature 30±2°C, humidity 6...

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Abstract

The invention provides an insect chitin synthase 1B gene segment, dsRNA and application thereof. The dsRNA is synthesized by the chitin synthase 1B gene segment with the sequence of SEQ ID NO:1 which is obtained by the nucleotide sequence of the chitin synthase 1B of Locusta migratoria manilensis with the accession number of GU067731 in an NCBI database. After the dsRNA is injected in body cavities of insects, the insects die because of difficult moulting. A new way for developing a safe and pollution-free method for preventing and controlling insect is provided.

Description

technical field [0001] The present invention relates to the field of biotechnology. It specifically relates to an insect chitin synthase 1B gene fragment and its dsRNA and the application of the dsRNA in killing pests. Background technique [0002] The damage caused by agricultural pests is an important factor restricting agricultural production in my country. Long-term application of chemical insecticides leads to a series of problems: 1) Insects become resistant to insecticides, the dosage of pesticides increases, and the cost of control increases; 2) Pesticide residues lead to serious environmental pollution; 3) Has a greater impact on non-target organisms. Existing biopesticides are widely used in pest control, but the insecticidal time is long and the effect is slow. [0003] RNA interference (RNAi) is a specific post-transcriptional gene silencing phenomenon caused by double-stranded RNA molecules, which won the Nobel Prize in 2006. The discovery of RNAi not only pro...

Claims

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Application Information

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IPC IPC(8): C12N15/52C12N15/113A01N61/00A01P7/04
Inventor 刘晓健张建珍杨美玲马恩波郭亚平
Owner SHANXI UNIV
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