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Paddy hpfr gene, expression product and application thereof

A technology for expressing products and rice, which can be applied in the field of agricultural biology and can solve problems such as unreported application research.

Inactive Publication Date: 2009-06-03
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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  • Paddy hpfr gene, expression product and application thereof
  • Paddy hpfr gene, expression product and application thereof
  • Paddy hpfr gene, expression product and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] 1. Discovery and cloning of rice hpfr gene based on Xanthomonas oryzae bacterial blight pathogenic variety hrfA Xoo Gene open reading frame design rice hpfr gene primers:

[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 the recombinant plasmid pETR[pET30a(+)::hpfr]( Figure 5 ). Transform pETR into Esch...

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] Cloning and expression of hpfw gene homologous to rice hpfr gene in wheat in embodiment 3

[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. Th...

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Abstract

The invention relats to rice hpfr gene, expression products and application thereof, belongs to biotechnology field for curepreventing plant diseases and insect pests and accelerating growthing and improving crop producibility. The invention is rice hpfr gene, which first discovering gene sequence with like function that of plant pathogenic bacteria hpfr gene in plant, coding product has exciton function and generating allergic reaction on tobacco plant, inducing palnt to generate disease resistance, accelerating vegetation and improving crop producibility, for the hpfr gene steming from plant, expression product hpfr protein can be generated by efficient expression carrier and microbe fermentation, which is a economical and environmental protection biological pesticide used in plowland and for curepreventing plant diseases and insect pests and improving crop producibility. The invention also provides wheat hpfw gene whose expression pfoduct hpfw protein having function of generating allergic reaction on tobacco plant, inducing palnt to generate disease resistance, accelerating vegetation.

Description

1. Technical field [0001] 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 [0002] 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 various environmental stress resistance capabilities. [0003] The hrp gene of plant pathogenic bacteria has been studied in detail at home and abroad. Wei et al. first discovered that a new protein en...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/82
Inventor 邵敏高学文黄万春郭玲王金生
Owner NANJING AGRICULTURAL UNIVERSITY
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