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Rice brown planthopper resistance gene Bph6 and molecular markers closely linked with same

An anti-BPH and molecular marker technology, applied in the field of plant genetic engineering, can solve the problems of difficult operation, limited effect, increased production cost, etc., and achieve clear function and good effect

Active Publication Date: 2016-11-23
WUHAN UNIV
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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 Bph6 and molecular markers closely linked with same
  • Rice brown planthopper resistance gene Bph6 and molecular markers closely linked with same
  • Rice brown planthopper resistance gene Bph6 and molecular markers closely linked with same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Example 1 Positional cloning of Bph6 gene and development of linked molecular markers

[0068] 1. Preliminary positioning results of Bph6

[0069] A Bph6-containing F 2 Populations, 93-11 and Swarnalata were all obtained from the National Crop Germplasm Conservation Center of the Institute of Crop Science, Chinese Academy of Agricultural Sciences, and used the CTAB method (Murray MG&Thompson, 1980 Rapidisolation of high-molecular-weight plant DNA.Nucleic Acids Res 8:4321-4325) Extract parent and F 2 Genomic DNA of each individual plant in the population. each F 2 A single plant obtains the corresponding F 2:3 family lineage. To identify F 2 The N. lugens resistance phenotype of each individual plant in the localization population was investigated by using the seedling stage group method to investigate the resistance performance of each individual plant of the F2:3 family. 2:3 Family resistance level represents F 2 BPH-resistant phenotype of individual plants. T...

Embodiment 2

[0092] Example 2 Functional Verification and Application of Bph6 Gene

[0093] 1. Construction of genetic transformation vector

[0094] (1) Construction of Bph6 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.

[0095] 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 Bph6 gene overexpression vector, which is electrotransformed 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.

[0096] (2) Bph6 gene RNAi vector construction

[0097] The vector used is pCXUN (provided by Professor Wang Guoliang of Ohio ...

Embodiment 3

[0103] Example 3 Verification of molecular markers

[0104] 1. Materials and methods

[0105] 1.1 Materials: BPH-resistant parent Swarnalata (containing the brown planthopper-resistant gene Bph6), brown planthopper-susceptible rice variety Yangdao 6 (93-11) and F produced by crossing Swarnalata and Yangdao 6 23 family lineage.

[0106] Molecular marker primers: H and Y37, the nucleotide sequences of which are respectively shown in SEQ ID No.4-5.

[0107] 1.2 Method

[0108] Genomic DNA was extracted from rice samples by CTAB extraction. Sample DNA was amplified with primers H and Y37, respectively. 10 μl system. The 10 μl reaction system includes: 10×PCR buffer, 1.0 μl; 10 mM dNTPs, 0.1 μl; 10 μM primer, 0.4 μl; 5U / μl Taq DNA polymerase, 0.2 μl and 50ng DNA template. The amplification reaction was carried out on a Bioer PCR instrument: 94°C for 4min; 30 cycles of 94°C for 30s, 55°C for 30s, 72°C for 90s; 72°C for 5min. The amplified products of H were separated by 1% ag...

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Abstract

The invention provides a rice brown planthopper resistance gene Bph6 and molecular markers closely linked with the same. A nucleotide sequence of the rice brown planthopper resistance gene Bph6 is represented as SEQ ID No.1, and a cDNA (complementary desoxyribonucleic acid) sequence of the rice brown planthopper resistance gene Bph6 is represented as SEQ ID No.2; the gene Bph6 is positioned between a molecular marker H and a molecular marker Y37 and are closely linked with the two molecular markers; rice containing the brown planthopper resistance gene Bph6 can be screened by the molecular marker H and the molecular marker Y37. The gene Bph6 is transferred into a common rice variety through genetic transformation and hybridization, and resistance of the rice to brown planthopper can be improved, so that damage caused by the brown planthopper is reduced and yield increase and stable yield are realized.

Description

technical field [0001] The invention belongs to the field of plant genetic engineering, in particular to a rice brown planthopper resistance gene Bph6 and its closely linked molecular markers. Background technique [0002] Rice is an important food crop, which is the staple food for more than half of the world's people. At the same time, 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 faced by people all over th...

Claims

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Application Information

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IPC IPC(8): C12N15/29C07K14/415C12N15/82C12Q1/68C12N15/11
CPCC07K14/415C12Q1/68C12N15/11C12N15/63C12N15/82A01H1/045
Inventor 何光存郭建平杜波邱永福陈荣智
Owner WUHAN UNIV
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