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Rice brown planthopper resistance gene bph37, protein, vector, host cell, molecular marker, method and application

A brown planthopper resistance and molecular marker technology, applied in application, genetic engineering, plant genetic improvement and other directions, can solve the problems of non-target biological poisoning, increased drug resistance of brown planthopper, increased production cost, etc., to reduce damage and improve brown planthopper resistance. Effect

Active Publication Date: 2021-10-22
WUHAN UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since the brown planthopper outbreaks mostly occur in the mature rice filling stage, when the rice plants are growing vigorously, it is very difficult to apply insecticides to the base of the rice plants.
In fact, due to the large-scale application of chemical insecticides year after year, the resistance of brown planthoppers has doubled, making the effect of pesticide control limited.
At the same time, the use of chemical pesticides to control brown planthoppers, on the one hand, increases the production cost of farmers, and on the other hand, chemical pesticides also cause environmental and ecological problems such as poisoning of non-target organisms, environmental and food pollution, etc.

Method used

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  • Rice brown planthopper resistance gene bph37, protein, vector, host cell, molecular marker, method and application
  • Rice brown planthopper resistance gene bph37, protein, vector, host cell, molecular marker, method and application
  • Rice brown planthopper resistance gene bph37, protein, vector, host cell, molecular marker, method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0060] Example 1 Cloning of Bph37 gene and development of molecular markers

[0061] 1. GWAS

[0062] Obtained 1520 rice varieties from the Institute of Crop Science, Chinese Academy of Agricultural Sciences, covering 12 types, located in 74 countries around the world ( figure 1 ). 1520 rice varieties were tested for resistance to brown planthopper by measuring the body weight of female adults of brown planthopper before and after feeding. Eight seeds of each test rice variety were sown and grown in a plastic cup (9 x 15 cm). A resistance test was carried out on rice plants at the five-leaf stage. Freshly emerged N. lugens biotype I female adults were collected and weighed with an electronic balance (Shimadzu; type: AUW120D). The brown planthopper with initial body weight of 1.80-2.70 mg was selected for experiment. Each N. lugens was put into a 2 x 2 cm film bag made of parafilm, and then fixed on a rice plant 1 cm away from the soil. The N. lugens fed on the leaf sheat...

Embodiment 2

[0073] Example 2 Functional Verification and Application of Bph37 Gene

[0074] 1. Construction of genetic transformation vector

[0075] Construction of Bph37 gene overexpression vector. The vector used is pCXUN (provided by Professor Wang Guoliang of Ohio State University in the United States). The pCXUN vector is cut with XcmI, and the foreign fragment can be directly connected after adding A.

[0076] According to the results of RACE, PCR method was used to amplify the ORF directly, and after adding A, it was connected into the vector. After the sequence verification is correct, the obtained vector is the Bph37 gene overexpression vector, which is transformed into Agrobacterium EHA105. Pick a single clone for expansion and culture, and after PCR verification, add an equal volume of 50% glycerol to mix, and store at -70°C for later use.

[0077] 2. Genetic Transformation

[0078] The genetic transformation method mediated by Agrobacterium EHA105 (Hiei et al., 1994, Effici...

Embodiment 3

[0081] Example 3 Verification of Molecular Markers

[0082] 1. Materials and methods

[0083] 1.1 Materials: BPH-resistant parent SE382 (containing rice BPH-resistant gene Bph37), brown planthopper-susceptible rice variety Yangdao 6 (93-11), and F hybridized between SE382 and Yangdao 6 2 strain.

[0084] Molecular marker primers: 156-35, the nucleotide sequences of which are respectively shown in SEQ ID No.4-5.

[0085] 1.2 Method

[0086] Genomic DNA was extracted from rice samples by CTAB extraction. Sample DNA was amplified with primer 156-35. 10 μl system. 10μl reaction system includes: 2×Es Taq MasterMix (Dye), 5.0μl; ddH 2 O, 3.7 μl; 10 μM primer, 0.4 μl; and 50 ng DNA template. The amplification reaction was carried out on a BioerPCR instrument: 94°C for 3min; 94°C for 30s, 55°C for 30s, 72°C for 60s, 30 cycles; 72°C for 5min. The amplified products of 156-35 were separated by 1% agarose gel and analyzed directly after electrophoresis.

[0087] 2. Results: Usin...

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Abstract

The invention discloses a rice brown planthopper resistance gene Bph37, protein, carrier, host cell, molecular marker, method and application. The nucleotide sequence of the rice brown planthopper resistance gene Bph37 is shown in SEQ ID No.1, and its cDNA sequence is shown in SEQ ID No.2. Using the molecular marker 156‑35, the rice containing the brown planthopper resistance gene Bph37 can be screened. Through genetic transformation, the Bph37 gene is transferred into ordinary rice varieties, which can improve the resistance of rice to brown planthoppers, thereby reducing the damage caused by brown planthoppers, and achieving the purpose of increasing and stabilizing yields.

Description

technical field [0001] The invention belongs to the field of plant genetic engineering, and specifically relates to a rice brown planthopper resistance gene Bph37, protein, carrier, host cell, molecular marker, method and application. Background technique [0002] Rice is an important food crop, which is the staple food for more than half of the world's people. Since the fine genetic map and physical map of the rice genome have been completed, its transgenic technology is relatively easy, and it has collinearity with other grass crop genomes, so it is regarded as a model plant. With the completion of genome sequencing of various organisms including rice, human beings have entered the post-genome era. It has become the frontier field of life science to carry out functional genome research in an all-round way. Therefore, the study of rice functional genes is of great significance to socioeconomic development and biological research. [0003] Food security is a challenge fac...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/82A01H5/00A01H6/46C12Q1/6895C12N15/11
CPCC07K14/415C12N15/8286C12Q1/6895C12Q2600/13
Inventor 何光存周聪杜波陈荣智祝莉莉
Owner WUHAN UNIV
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