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Wheat hpfw gene, expression product and uses thereof

A wheat and gene technology, applied in the field of agricultural biology, can solve problems such as unreported applied research

Inactive Publication Date: 2010-11-03
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, the hrp gene and its application in plant pathogenic bacteria have been studied more, but the harpin-encoding gene and its application in plants have not been reported so far

Method used

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  • Wheat hpfw gene, expression product and uses thereof
  • Wheat hpfw gene, expression product and uses thereof
  • Wheat hpfw gene, expression product and uses thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] 1. Discovery and cloning of rice hpfr gene The rice hpfr gene primers were designed according to the open reading frame of Xanthomonas oryzae bacterial blight pathogenic variety hrfAXoo gene:

[0047] 5'-GGAATTCCATATGAACTCTTTGAACACACAATTGG-3' and 5'-CGGGATCCTATTACTGCATTGATGCGCTTCC-3' were used to amplify the hpfr gene from rice by PCR method. PCR amplification conditions: 1 cycle of 94°C / 5min, 35 cycles of 94°C / 45sec+60°C / 45sec+72°C / 45sec, 1 cycle of 72°C / 7min. The gene fragment was named hpfr gene ( figure 1 ). The rice hpfr gene was used as a probe to hybridize with the total DNA of the rice genome. The hybridization results showed that there was a hybridization positive signal in the rice genome ( figure 2 ).

[0048] 2. In vitro expression of rice hpfr gene

[0049] After the PCR product was purified, it was digested with NdeI and BamHI, and connected to the expression vector pET30a(+) to obtain a recombinant plasmid pETR[pET30a(+)::hpfr]( Figure 5 ). Transf...

Embodiment 2

[0052] Example 2 Direct use of rice hpfr gene expression product hpfr protein

[0053] The rice hpfr gene expression product hpfr protein is sprayed on the leaves of the plants at the concentration of 15, 30, and 60 μg / ml.

[0054] 1. Inducing the resistance phenotype of Xanthi tobacco to TMV

[0055] Combine rice hpfr and harpin Xoo The protein CFEP mother liquor was diluted to concentrations of 15, 30, and 60 μg / ml, respectively, and sprayed with a micro-sprayer on Shanxi tobacco at the adult plant stage, and each treatment was repeated 5 times; 16 hours later, the activated TMV was inoculated on the tobacco leaves by friction; 4 Days later, observe the results and count the number of lesions. The results of the study showed that rice hpfr and harpin Xoo Treatment of tobacco, all have the ability to obviously induce tobacco resistance to TMV (decrease in the number of scabs) ( Figure 8 ). Multiple comparison analysis by LSD method (F 189.464 ) showed that BL21(DE3) / pE...

Embodiment 3

[0063] Example 3 Cloning and expression of hpfw gene homologous to rice hpfr gene in wheat

[0064] 1. Design primers for wheat hpfw gene cloning and nucleotide sequence:

[0065] 5'-GGAATTCCATATGAACTCTTTGAACACACAATTGG-3' and 5'-CGGGATCCTATTACTGCATTGATGCGCTTCC-3' were used to amplify the homologous gene of rice hpfr gene from wheat and named as hpfw gene by PCR method (the same method as in Example 1). The nucleotide sequence of the hpfw gene is SEQ ID NO.3 ( Figure 11 ), the size is 414bp, and the homology with the rice hpfr gene is 93%. The deduced amino acid sequence of hpfw gene is SEQ ID NO.4( Figure 12 ).

[0066] 2. In vitro expression of wheat hpfw gene and small quantity production hpfw protein (method is the same as embodiment 1)

[0067] The wheat hpfw gene was connected to the expression vector pET30a(+) to obtain the recombinant expression vector pETW9[pET30a(+)::hpfw], and the product expressed by Escherichia coli BL21(DE3) was named hpfw protein. This thr...

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Abstract

The invention relates to a wheat hpfw gene, an expression product and application thereof which belong to the technical field of agricultural biology, and are used for preventing plant disease and pests, improving plant growth and crop yield. The rice hpfr gene has a gene sequence with functions similar to those of plant pathogenetic bacteria hrp gene in plant, which is discovered for the first time; and a coded product has functions of exciton, such as generating anaphylactic reaction on tobacco, inducing plant to generate disease resistance, improving plant growth, improving crop yield and the like; as the hpfr gene originates from plant, and the expression product hpfr protein is generated through expressing vector with high efficiency and microbial fermentation, the gene and the expression product are economic and environment-friendly biological pesticide, and can be used for preventing and treating plant disease and pests in land and sheltered land and improving the yield. The invention also provides the wheat hpfw gene, wherein an expression product hpfw protein of the gene has the functions of inducing tobacco to generate anaphylactic reaction, exciting plant to generate resistance, and promoting plant growth.

Description

[0001] This application is a divisional application with application date: February 14, 2007, application number: 200710020342.X, name: rice hpfr gene, expression product and application thereof. 1. Technical field [0002] The invention relates to rice hpfr gene, expression product and application thereof, belongs to the field of agricultural biotechnology, and is used for preventing and treating plant diseases and insect pests, promoting plant growth and increasing crop yield. 2. Background technology [0003] The hrp gene is a gene in plant pathogenic bacteria that determines the hypersensitive response (hypersensitive response, HR) on non-host plants and pathogenicity on host plants [Lindgren P B, The role of hrp genes during plant-bacterial interaction.Annu Rev Phytopathol, 1997 , 35: 129-152]. Anaphylaxis is a localized and rapid form of programmed cell death (PCD) produced by plants. PCD can trigger one or more signal transduction pathways, endowing plants with vario...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/10C07K14/415A01N63/00A01P3/00A01P21/00
CPCY02A40/10
Inventor 邵敏郭玲高学文
Owner NANJING AGRICULTURAL UNIVERSITY
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