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Cytochrome P450 dsRNA (double-stranded ribonucleic acid) and application to aphid growth inhibition

A cytochrome and aphid technology, applied in the direction of DNA / RNA fragments, applications, plant growth regulators, etc., can solve problems such as economic losses, conventional breeding is difficult to work, and resistance mechanisms are not clear

Active Publication Date: 2012-11-14
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

[0004] There is a lack of aphid-resistant germplasm resources in wheat and vegetables, and the resistance mechanism is still unclear. Conventional breeding is difficult to work, causing huge economic losses to agricultural production every year

Method used

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  • Cytochrome P450 dsRNA (double-stranded ribonucleic acid) and application to aphid growth inhibition
  • Cytochrome P450 dsRNA (double-stranded ribonucleic acid) and application to aphid growth inhibition
  • Cytochrome P450 dsRNA (double-stranded ribonucleic acid) and application to aphid growth inhibition

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Experimental program
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Embodiment 1

[0038] Embodiment 1, the acquisition of cytochrome P450dsRNA

[0039] 1. Extraction of total RNA and synthesis of cDNA

[0040] Take about 20 tube aphids and peach aphids respectively, extract total RNA according to the instructions of the Trizol kit of Beijing Quanshijin Company, purify it with RNACleaning Kit, and synthesize the first strand of cDNA by reverse transcription. The operation steps refer to the instructions of the kits.

[0041] 2. Primer design and gene cloning

[0042]According to the sequences of the cytochrome P450 of pea aphid and cotton aphid (GenBank accession numbers are NM_001163211.2 and JQ246416.1, respectively), DNAman5.0 was used for alignment, and the conserved sequence was selected, and Primer Primer5.0 software was used to design primer P1 (Table 1 ), 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...

Embodiment 2

[0060] Example 2, the application of dsRNA (P450dsRNA) in inhibiting the growth of aphids

[0061] 1. Preparation of artificial diet for aphids and preparation of feeder

[0062] 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.

[0063] 2. Application of dsRNA (P450dsRNA) in inhibiting the growth of aphids

[0064] Aphid feeding methods refer to the following documents: Jiao Min, Liu Shusheng. The technology of raising aphids wi...

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Abstract

The invention discloses dsRNA (double-stranded ribonucleic acid) and application to aphid growth inhibition. The dsRNA provided by the invention is dsRNA as expressed by 1) or 2) as follows: 1) dsRNA consisting of ribonucleic acid as shown by a sequence 4 in a sequence table and ribonucleic acid as shown by a reverse complementary sequence of the sequence 4; and 2) dsRNA consisting of ribonucleicacid as shown by a sequence 5 in the sequence table and ribonucleic acid as shown by a reverse complementary sequence of the sequence 5. According to experiments, the obtained dsRNA of a conserved sequence of cytochrome P450cDNA (complementary deoxyribonucleic acid) of English grain aphids and green peach aphids can inhibit growth and development of the English grain aphids and the green peach aphids and can cause a lethal effect by adopting a method of feeding the dsRNA in vitro and using an RNAi (ribonucleic acid interfere) technology to silence the in-vivo cytochrome p450 of the English grain aphids and the green peach aphids.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a cytochrome P450 gene dsRNA and its application in inhibiting the growth of aphids. Background technique [0002] 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. Peach aphid is a wide-feeding pest with about 350 kinds of hosts. It mainly harms tobacco, peach, plum, plum, pear and other fruit trees and vegetables such as cabbage, radish, pepper and spinach, causing great economic losses. 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 in...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/63C12N5/10C12N1/21A01N57/16A01N63/00A01P7/04
Inventor 夏兰琴王晖张岷
Owner INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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