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DsRNA of gene of symbiotic bacterium of aphids and application thereof to inhibition of growth of aphids

A technology for symbiotic bacteria and aphids, applied in DNA/RNA fragments, applications, recombinant DNA technology, etc.

Active Publication Date: 2014-10-22
INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The research on the indirect control of aphids by silencing symbiont-related genes through RNAi technology has not yet been reported

Method used

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  • DsRNA of gene of symbiotic bacterium of aphids and application thereof to inhibition of growth of aphids
  • DsRNA of gene of symbiotic bacterium of aphids and application thereof to inhibition of growth of aphids
  • DsRNA of gene of symbiotic bacterium of aphids and application thereof to inhibition of growth of aphids

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1. Obtaining the dsRNA of the aphid endosymbiont Hamiltonella defensa28469 gene

[0027] 1. Extraction of wheat aphid total RNA and synthesis of cDNA

[0028] About 20 wheat aphids were taken respectively, and total RNA was extracted according to the instructions of the Trizol kit of Beijing Quanshijin Company, purified with RNA Cleaning Kit, and the first strand of cDNA was synthesized by reverse transcription. The operation steps refer to the instructions of the kit.

[0029] 2. Primer design and gene cloning

[0030] The aphid symbiont-related gene 28469 (a fragment of 28469 was used for RNAi interference) was obtained by sequencing the intestinal transcriptome of wheat aphid. Primer P1 (Table 1) was designed using Primer Primer5.0 software and synthesized by Beijing Huada Gene Company. The green fluorescent protein gene (GFP) fragment was amplified from the GFP plasmid preserved in the laboratory of the National Wheat Improvement Center, and primer P2 was d...

Embodiment 2、28469

[0046] Example 2, the application of 28469dsRNA in inhibiting the growth of aphids

[0047] 1. Preparation of artificial diet for aphids and preparation of feeder

[0048] References for the preparation of artificial feed and feeder (Li Caixia, Gao Lifeng, Gao Lingling, Li Runzhi. Research on feeding aphids in pure artificial nutrient solution. Journal of Shanxi Agricultural University, 1997, 17(3): 225-228. Li C X , Gao L F, Gao L L, Li R Z. Study on the rearing of aphids on an artificially solidic diets. Journal of Shanxi Agricultural University, 1997,17(3):225-228.(in Chinese)), with 0.2μm Filter the artificial feed with a bacterial filter, dispense it into 2.0mL sterilized centrifuge tubes and store in a -20°C refrigerator to avoid repeated freezing and thawing.

[0049] 2. Application of dsRNA (28469dsRNA) in inhibiting the growth of aphids

[0050] Aphid feeding methods refer to the following documents: Jiao Min, Liu Shusheng. The technology of raising aphids with arti...

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Abstract

The invention discloses dsRNA of a gene of a symbiotic bacterium of aphids and an application thereof to inhibition of growth of aphids. DsRNA provided by the invention is double-stranded RNA formed by a nucleotide shown in a sequence 2 in a sequence table and a nucleotide shown in an inverse complementary sequence of the sequence 2. Experiments prove that the gene of the endotrophic symbiotic bacterium hamiltonella defensa 28469 of wheat aphids is obtained; the gene of the endotrophic symbiotic bacterium hamiltonella defensa 28469 of wheat aphids can be inhibited or silenced, impaired growth and development of the wheat aphids can be caused and lethal effects can be generated by adopting the method of feeding dsRNA in vitro.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a dsRNA of aphid symbiotic bacteria gene and its application in inhibiting the growth of aphids. Background technique [0002] Wheat aphids are one of the main pests that harm wheat production in China. According to statistics, the area of ​​damage caused by wheat aphids in China can reach as high as 17 million hectares every year, accounting for 62% of the total wheat planting area, resulting in a 15%-30% reduction in production, and even up to 50% in severe cases. %. In recent years, due to factors such as global warming and changes in farming systems, the reproductive ability and adaptability of aphids have been significantly enhanced, and their harm has become increasingly serious. At present, aphid control is mainly based on spraying pesticides, but a large amount of pesticides are not only harmful to humans and animals, but also cause serious environmental pollution. Breeding...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/11A01N57/16A01P7/04
Inventor 夏兰琴孙永伟蔡洵
Owner INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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