Application of BZR1 gene in regulation and control of insect attack stress resistance of plants

A gene and insect pest technology, applied in the fields of application, plant peptides, plant products, etc., can solve the problems of unclear function and mechanism of BZR1, and achieve the effect of enhancing resistance

Active Publication Date: 2021-12-17
ZHEJIANG UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the role and mechanism of BZR1 in pest stress are still unclear

Method used

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  • Application of BZR1 gene in regulation and control of insect attack stress resistance of plants
  • Application of BZR1 gene in regulation and control of insect attack stress resistance of plants
  • Application of BZR1 gene in regulation and control of insect attack stress resistance of plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1 Preparation and identification of tomato SlBZR1 deletion mutant plants

[0042] 1. Construction of CRISPR / Cas9 vector containing specific sgRNA

[0043] First, find the full-length DNA sequence of the tomato SlBZR1 gene on the SGN website https: / / solgenomics.net / , as shown in SEQ ID NO.4, and input it into the http: / / crispr.hzau.edu.cn / CRISPR2 / website , Find out the 20bp base sequence before the PAM structure of the protein coding region with a high onscore score and GC content >40%, as the sgRNA specifically targeting the protein coding region of the gene, as shown in SEQ ID NO.5.

[0044] The single-stranded sgRNA sequence was synthesized and then annealed to form a double-stranded sgRNA with BbsI restriction endonuclease sites at both ends. The above synthetic fragment was inserted into the BbsI site of the AtU6-sgRNA-AtUBQ-Cas9 vector to obtain an intermediate vector.

[0045] Using HindⅢ and KpnⅠ restriction endonucleases to carry out double enzyme dige...

Embodiment 2

[0055] Example 2 Identification of tomato SlBZR1 gene resistance to cotton bollworm phenotype

[0056] Cotton bollworms were 3rd instar cotton bollworms purchased by Henan Jiyuan Baiyun Industrial Co., Ltd., and cotton bollworms of the same size were selected into 24-well plates, and starved for one night.

[0057] When the tomato seedlings grow to six leaves and one heart, select the leaves at the 2-4 nodes from top to bottom, weigh the leaves with the same mass (1±0.05g) and place them in a petri dish with 1% agar to prevent in vitro Leaves were dehydrated, and the tomato leaves before inoculation were photographed and recorded. Then two starvation-treated cotton bollworms were inoculated on the tomato leaves of a petri dish, and the petri dish was covered and fixed with rubber bands to prevent the bollworms from leaving the petri dish. After 3 days of inoculation treatment under low light, the weight of the cotton bollworm was detected, and then the cotton bollworm was soa...

Embodiment 3

[0059] Example 3 Effect of Tomato SlBZR1 Gene on Resistance Hormone JA

[0060] When the tomato seedlings grow to six leaves and one heart, select the leaves at the 3-4 nodes from top to bottom for W+OS simulated cotton bollworm feeding treatment, and collect leaf samples at 0 and 1 hour to analyze the JA hormone level, each time point Set up 4 repetitions.

[0061] W+OS simulated cotton bollworm feeding as follows: use hemostatic forceps to clamp the leaves on both sides of the main vein of the leaflet to create mechanical wounds, and then apply five times diluted cotton bollworm mouthpart secretions to the wounds.

[0062] The method of obtaining the mouthpart secretions of the cotton bollworm is as follows: after the cotton bollworms are starved for 12 hours, they are fed tomato leaves, and after 12 hours, the mouthpart secretions of the cotton bollworms are absorbed as a stock solution.

[0063] JA hormone determination method: Weigh 0.1g tomato leaves, grind with liquid ...

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Abstract

The invention discloses application of a BZR1 gene in regulation and control of insect attack stress resistance of plants, and belongs to the technical field of biology. According to the application, the function of the BZR1 gene is deleted by utilizing a biotechnological means, so that the resistance of plants to insect attack stress is improved; and the coding sequence of the BZR1 gene is as shown in SEQ ID NO.1 or has at least 70% of homology with the sequence as shown in SEQ ID NO.1, and the coded protein is equivalent in function. The invention discloses the application of the BZR1 gene in regulating and controlling the insect attack stress, especially the helicoverpa armigera resistance of the plant for the first time, the function of the BZR1 gene is deleted through a gene editing technology, the helicoverpa armigera resistance of the plant can be remarkably enhanced, and the BZR1 gene can be applied to cultivation of insect attack stress resistant plant varieties.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for improving plant resistance to insect pests, especially cotton bollworm, by deleting BZR1 gene function. Background technique [0002] Tomato (Solanum lycopersicum L.) is one of the most widely consumed and grown vegetables in the world. However, during the whole growth cycle, tomatoes are attacked by various diseases and insect pests, resulting in huge economic losses. If a large amount of pesticides are used for control, it will bring problems such as pesticide residues, environmental pollution and drug resistance. In view of the relatively short life cycle of tomato, small genome, high genetic diversity, easy transgenic and cultivation, coupled with the completion of whole genome sequencing and the development of molecular genetics research, tomato has become a classic model for studying the interaction between plants and pests. plant. Therefore, it is of positive e...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/29C12N15/82C12N15/113C07K14/415A01H5/00A01H6/82
CPCC07K14/415C12N15/8218C12N15/8286
Inventor 喻景权吴少芳胡璋健胡超轶范飞军周艳虹
Owner ZHEJIANG UNIV
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