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Damage-inducible promoter of rice stem borer and its application

A rice stem borer and promoter technology, applied in the field of plant genetic engineering, can solve the problems of destroying the natural regulation function of the rice field ecosystem, drug residues in rice plants, and mutual interference of lures.

Active Publication Date: 2021-06-15
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, chemical pesticides have become the first choice tool for pest control. However, the long-term and large-scale application of pesticides in rice fields has significantly destroyed the natural regulation function of rice field ecosystems, and various safety problems such as pesticide resistance of pests and drug residues in rice plants have also emerged. Come
Later, some scholars proposed to use sexual lures to trap or release natural enemies to complete the control of rice field pests. However, only one lure can be used for sex lures at a time, and multiple lures will interfere with each other. The method of releasing natural enemies There are problems in the selection of natural enemy populations, the cultivation of natural enemies and the release in the field

Method used

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  • Damage-inducible promoter of rice stem borer and its application
  • Damage-inducible promoter of rice stem borer and its application
  • Damage-inducible promoter of rice stem borer and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Example 1: The rice stem borer feeding-induced response promoter is the rice stem borer damage-inducible promoter P HPL2 and the acquisition of corresponding missing fragments

[0031] If the present invention has no special instructions, the reference method used and the basic operation of corresponding molecular biology have all referred to: J. Sambrook etc., " Molecular Cloning Experiment Guideline (Second Edition) " (Chinese translation), Science Press 1996 version information. Genomic DNA extraction of rice "Zhonghua 11": Take 3 cm of fresh leaves of rice at the tillering stage, and the method refers to the CTAB extraction method published by Murray in 1980 (Murray MG, Thompson WF. Rapid isolation of high molecular weightplant DNA. Nucleic Acids Res. 1980,8:4321-5), the extracted DNA was completely dissolved and stored in a -20°C refrigerator.

[0032] The upstream sequence of the start codon ATG of the OsHPL2 (LOC_Os02g12680) gene sequence was obtained from the ...

Embodiment 2

[0035] Embodiment 2: Construction of plant expression vector

[0036] The carrier DX2181 was double-digested with BamHI and SalI, and the vector fragment was recovered with the DNA gel recovery kit of Tiangen Biochemical Technology Co., Ltd., see the kit manual for specific operations, and the integrity of the enzyme digestion was detected by electrophoresis, and stored in a -20°C refrigerator.

[0037] Digest P with BamHI and SalI HPL2 And the TA cloning plasmid of the corresponding deletion fragment. Use the same method to recover the target fragment and check the integrity of the fragment. Store in -20°C refrigerator.

[0038] Recycled P HPL2 The fragment and the corresponding deletion fragment were constructed on the vector DX2181 through the cohesive end of the enzyme cutting site (for the vector map and multiple cloning sites, see figure 2 ), electrotransformed into Escherichia coli DH5α. After enzyme digestion and sequencing, the recombinant expression vector DX218...

Embodiment 3

[0042] Example 3: Agrobacterium-mediated genetic transformation

[0043] The Agrobacterium-mediated genetic transformation method of the present invention refers to the method described in the "Agrobacterium-mediated Genetic Transformation Operation Manual" published by the State Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University (Lin Yongjun et al., 2002). The embryogenic callus induced by the rice variety "Zhonghua 11" (from the Institute of Crop Science, Chinese Academy of Agricultural Sciences) was transformed. Respectively through callus induction culture, callus subculture, Agrobacterium infection, co-cultivation, screening to obtain hygromycin resistant resistant callus, and then through differentiation, rooting, seedling hardening and transplanting to obtain transgenic plants . The genetic transformation steps mediated by Agrobacterium involved in the present invention and the medium formula are as follows.

[0044] 1. Abbreviations of reage...

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Abstract

The invention discloses a damage-inducible promoter of rice stem borer and its application. The damage-inducible promoter of rice stem borer is named: P HPL2 , its length is 2009bp, and its nucleotide sequence is shown in SEQ ID NO:1. The promoter is the promoter of the lipid hydroperoxide lyase gene OsHPL2 (LOC_Os02g12680), which is the key gene for cloning and synthesizing green leaf volatiles GLVs from the rice "Zhonghua 11" genome, and has a positive effect on the promoter P HPL2 The deletion resulted in six deletion fragments of different lengths at the 5' end, and the study found that there were positive regulatory elements specifically expressed by the feeding of C. These seven promoters and their corresponding expression vector preparation methods, as well as the rice genetic transformation method mediated by Agrobacterium, the characteristics of the above promoters have potential application value in rice insect-resistant genetic engineering breeding.

Description

technical field [0001] The invention relates to the technical field of plant genetic engineering, in particular to a rice stem borer damage-inducible promoter and application thereof. Background technique [0002] During the whole growth process of rice, it suffers from various pests all the time. The most important pests are brown planthopper, rice stem borer and rice leaf roller. 10% of the annual yield loss of rice is caused by pests (Chen H, Lin Y, Zhang Q. Review and prospect of transgenic rice research. Chin Sci Bull. 2009, 54:4049–4068). Therefore, chemical pesticides have become the first choice tool for pest control. However, the long-term and large-scale application of pesticides in rice fields has significantly destroyed the natural regulation function of rice field ecosystems, and various safety problems such as pesticide resistance of pests and drug residues in rice plants have also emerged. Come. Later, some scholars proposed to use sexual lures to trap or r...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/113C12N15/82A01H5/00A01H6/46
CPCC12N9/88C12N15/8286
Inventor 华红霞李翰鹏蔡万伦
Owner HUAZHONG AGRI UNIV
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