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dsRNA (double-strand ribonucleic acid) and application of combination thereof to control aphid damage

A technology in aphids and sequence listings, applied in the direction of DNA/RNA fragments, applications, recombinant DNA technology, etc., can solve the problems of lack of aphid-resistant germplasm resources, unclear resistance mechanism, and difficulty in conventional breeding, etc.

Active Publication Date: 2014-02-12
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

[0005] There is a lack of aphid-resistant germplasm resources in wheat, and the resistance mechanism is still unclear. Conventional breeding is difficult to work, and it causes huge economic losses to agricultural production every year.

Method used

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  • dsRNA (double-strand ribonucleic acid) and application of combination thereof to control aphid damage
  • dsRNA (double-strand ribonucleic acid) and application of combination thereof to control aphid damage
  • dsRNA (double-strand ribonucleic acid) and application of combination thereof to control aphid damage

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Example 1. Acquisition of growth and development-related genes 8273dsRNA and 22544dsRNA

[0050] 1. Extraction of total RNA and synthesis of cDNA

[0051] About 20 tube 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.

[0052] 2. Primer design and gene cloning

[0053] According to the results of the transcriptome sequencing of the plant Aphid in our laboratory, the growth and development-related genes 8273 (sequence 1) and 22544 (sequence 2) were obtained, and primers P1 and P2 (Table 1) were designed by using Primer Primer5.0 software, provided by Beijing Hua Synthesized by Big Genomics. The green fluorescent protein gene (GFP) fragment was amplified from the GFP plasmid preserved in the laboratory of the National Wheat...

Embodiment 2

[0070] Embodiment 2, the application of dsRNA in inhibiting the growth of aphids

[0071] 1. Preparation of artificial diet for aphids and preparation of feeder

[0072] References for the preparation of artificial feed and feeder (Li Caixia, Gao Lifeng, Gao Lingling, Li Runzhi. Research on raising 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.

[0073] 2. Application of dsRNA (22544dsRNA, 8273dsRNA) in inhibiting the growth of aphids

[0074] Aphid feeding methods refer to the following documents: Jiao Min, Liu Shusheng. The technology of raising aphids...

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Abstract

The invention discloses a dsRNA (double-strand ribonucleic acid) and an application of combinations thereof to control aphid damage. The dsRNA consists of (1) a nucleotide shown as a sequence 4 in a sequence table and a nucleotide shown as a reversely complementary sequence of the sequence 4; (2) a nucleotide shown in a sequence 5 in the sequence table and a nucleotide shown as a reversely complementary sequence of the sequence 5; or (3) a dsRNA shown by (1) and a dsRNA shown by (2). Experiments show that effects of inhibiting the growth and development of sitobion avenae and causing the death of the sitobion avenae are achieved by feeding the dsRNA for growth and development related genes 8273 and 22544 cDNA (complementary deoxyribonucleic acid) of the sitobion avenae in vitro and silencing the growth and development related genes 8273 and 22544 of the sitobion avenae by utilizing an RNAi (RNA interference) technology.

Description

[0001] The application is based on the application date of January 9, 2013, the application number is 201310007173.1, and the invention name is "dsRNA A divisional application of the patent application of "Application of " and its combination in the control of aphid damage" technical field [0002] The invention relates to the field of biotechnology, in particular to the application of dsRNA and its combination in controlling aphid damage. Background technique [0003] Wheat aphids (especially wheat aphids) are one of the main pests that harm wheat production in China. According to statistics, the area of ​​wheat aphid damage 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% reduction in production— 30%, 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 significantl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113A01N57/16A01P7/04
Inventor 夏兰琴张珉李雪峰
Owner INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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