Preparation method and application of RNA interference sequence of trehalase gene of Acyrthosiphon pisum

A technology of RNA interference and trehalase, applied in the field of RNA interference sequence preparation

Active Publication Date: 2019-06-07
GANSU AGRI UNIV
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  • Claims
  • Application Information

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Problems solved by technology

The application of RNAi technology in pests is still in its infancy, and there are many

Method used

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  • Preparation method and application of RNA interference sequence of trehalase gene of Acyrthosiphon pisum
  • Preparation method and application of RNA interference sequence of trehalase gene of Acyrthosiphon pisum
  • Preparation method and application of RNA interference sequence of trehalase gene of Acyrthosiphon pisum

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Embodiment 1

[0031] Example 1: Preparation and application detection of an RNA interference sequence of the pea aphid trehalase gene

[0032] Experimental steps:

[0033] 1 dsRNA design and synthesis

[0034] According to the sequences submitted by 3 major international databases NCBI, trehalase (XM_003245847.3) was compared with MEGA software to deduce the conserved region, and the software Primer 5.0 was used to design specific amplification primers to amplify pea aphid trehalose synthase and seaweed Carbohydrase Conserved Region Gene. Send the amplified product to Xi'an Qingke for sequencing, and compare the gene sequencing results. The primer sequence is as follows: sense strand 5'-3'GGCGTGGTTTGACTGGGATA

[0035] Antisense strand 5'-3'CAACCAAAACCTGTCTGCGG

[0036] 2. Effects of feeding dsRNA on the mortality rate, gene silencing level, life table parameters, feeding behavior and trehalase activity of two color types of pea aphids

[0037] The concentration of the synthesized dsRNA ...

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Abstract

The invention discloses a preparation method of an RNA interference sequence of a trehalase gene of Acyrthosiphon pisum. The method comprises the following steps: 1. selecting a target region of RNAiaccording to a trehalase sequence of the Acyrthosiphon pisum (XM_003245847.3), and using Primer5.0 to design a PCR amplification primer in the target region; 2. extracting the total RNA of the Acyrthosiphon pisum and obtaining a cDNA by reverse transcription, and then conducting PCR-amplification on the trehalase gene contained in the cDNA by using the amplification primer to obtain the target gene; and 3. amplifying a double-stranded DNA of interest containing a T7promoter sequence, and then synthesizing a double-stranded RNA using the kit.

Description

technical field [0001] The application belongs to the field of pest control, and specifically relates to a preparation method and application of an RNA interference sequence of a pea aphid trehalase gene. Background technique [0002] The pea aphid (Acyrthosiphon pisum) belongs to the family Aphididae in the order Homoptera. It is widely distributed all over the world and is one of the main pests of many leguminous crops and pastures. In the alfalfa growing areas in Northwest my country, the pea aphid can cause economic losses of 10% to 30% of alfalfa production each year. When the pea aphid harms plants, it uses the piercing-sucking mouthparts to suck the phloem juice of the plant, thereby affecting the growth and development of the plant, normal life activities such as flowering and fruiting, and causing the death of the entire plant when the damage is serious. The pea aphid can be a carrier of plant viruses, and can transmit 25 kinds of viruses such as alfalfa mosaic vir...

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Application Information

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IPC IPC(8): C12N15/113C12N15/10A01N57/16A01P7/04
CPCY02A50/30
Inventor 王广张克信张艳蕾刘长仲
Owner GANSU AGRI UNIV
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