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Phytophthora capsici pectin methylesterase Pcpme l gene as well as preparation method of protein and application thereof

A technology of pectin methylesterase and Phytophthora capsici, applied in the field of molecular biology, can solve the problem that the target gene function of Phytophthora capsicum pectin methylesterase has not been published or published.

Inactive Publication Date: 2010-03-17
SHANDONG AGRICULTURAL UNIVERSITY
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Problems solved by technology

In the prior art, there is no disclosure or publication of the role of the PME target gene of Phytophthora capsici in the process of pathogen-host interaction

Method used

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  • Phytophthora capsici pectin methylesterase Pcpme l gene as well as preparation method of protein and application thereof
  • Phytophthora capsici pectin methylesterase Pcpme l gene as well as preparation method of protein and application thereof
  • Phytophthora capsici pectin methylesterase Pcpme l gene as well as preparation method of protein and application thereof

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Embodiment 1

[0055] Embodiment 1 (joint test Phytophthora capsici genomic DNA library construction)

[0056] Phytophthora capsici strains with strong pathogenicity and high PMEs activity were selected as test materials to construct a genomic DNA library. The specific steps are as follows:

[0057] (1) Using the CTAB method to extract high-quality DNA from the Phytophthora capsici genome;

[0058] (2) Ultrasonic fragmentation of genomic DNA. After fragmentation of genomic DNA, the 1.5Kb-3.0Kb DNA fragment obtained by electrophoresis detection was recovered and purified using the QIAEXII GEL Extraction Kit kit;

[0059] (3) Genomic library identification, after DNA fragments of appropriate size are connected to the vector, transform Escherichia coli DH5α, take 20 μL of the bacterial liquid and smear the LB plate containing Amp, X-gal and IPTG, and blue-white screening;

[0060] (4) 96 clones were randomly selected for colony PCR identification and sequencing analysis to ensure that more tha...

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Abstract

The invention provides a pectin methylesterase gene Pcpme l cloned from phytophthora capsici and a preparation technique of a protein thereof, belonging to the technical field of biology. The invention proves that the pectin methylesterase gene Pcpme l effectively participates in the processes of infecting a host by the phytophthora capsici and causing the disease course of phytophthora capsici leonian on the basis of gene and protein levels and also proves that the protein coded by the pectin methylesterase gene Pcpme l has property of destroying leaf tissue cells and cell wall structures thereof to enable destroyed parts to generate obvious symptoms on the basis of a cell chemistry technique, thus the pectin methylesterase gene Pcpme l is an important target pathogenic gene of a coded phytophthora capsici pectin methylesterase gene cluster. The invention provides sufficient technical reserve for further researching a germ molecule detection technique.

Description

technical field [0001] The invention belongs to the field of molecular biology. Specifically, the invention relates to a method for isolating the gene of Phytophthora capsici pectin methylesterase and preparing the encoded protein. In addition, the present invention also relates to the destructive effect of the pectin methylesterase encoded by the gene on the capsicum host. Background technique [0002] Phytopathogenic oomycetes secrete a variety of important cell wall degrading enzymes or pathogenic enzymes during the interaction process with the host, among which pectin methylesterase (PME) is an important pathogenic enzyme secreted by plant pathogenic oomycetes, which is produced in the pathogenic bacteria In the process of identification, colonization and interaction with the host, the invasion and colonization of pathogenic bacteria are promoted by degrading or softening the pectin of the cell wall, the mesoplast and its pectin aggregates. Therefore, pectin methylester...

Claims

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Application Information

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IPC IPC(8): C12N15/55C12N9/16C12P19/34C12N15/81C12Q1/44C12R1/84C12R1/645
Inventor 张修国
Owner SHANDONG AGRICULTURAL UNIVERSITY
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