Multiple PCR detection method for sugar-cane mosaic disease and ratoon stunting disease pathogeny

A detection method, mosaic disease technology, applied in biochemical equipment and methods, microbial measurement/inspection, organic chemistry, etc., can solve the problems of cumbersome detection and high detection cost, save time, simple operation, and save detection costs Effect

Inactive Publication Date: 2008-05-28
广西康田农业科技股份有限公司
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Problems solved by technology

This detection is more cumbersome and the detection cost is also high

Method used

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Embodiment Construction

[0012] The present invention will be described in further detail below.

[0013] The multiple PCR detection method of the sugarcane mosaic disease designed by the present invention and the pathogen of perennial dwarf disease comprises the following steps:

[0014] 1. Determine the best detection sampling site on the sugarcane plant:

[0015] Due to the respective distribution characteristics of SCMV and Lxx in sugarcane plants, in order to ensure that the two pathogens can be detected without omission in the materials to be tested, the sheathed leaves at the base of the stems of the sugarcane plants to be tested should be used as materials for extracting total nucleic acids.

[0016] 2. According to the known sugarcane mosaic virus dominant strain SCMV-A in mainland China and other four strains SCMV-B, D, E, SC (the sequence numbers in the GenBank database are U7354, U57355, U57356, U57357, D00948) nucleic acid conserved sequence region design reverse transcription primer (5'...

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Abstract

The invention discloses a multiple PCR detecting method for pathogen of sugarcane mosaic disease and ratoon stunting disease, which is characterized in that (1), a sheathing leaf at the stem base portion of a sugarcane plant is employed as detecting sample. (2), reverse transcription primer and primer pair of PCR detection cpc1 and cpc2 and RSD primer pair F1 and F2 are designed, sugarcane mosaic virus and genetic fragment of pathogen of sugarcane ratoon stunting disease are amplified. (3), total nucleic acid is extracted, and the RNA thereof is turned into reverse transcription via cDNA. (4), reaction system and amplification program of the one-step method with multiple RT-PCR are designed, and the nucleic acid is amplified. (5), gel electrophoresis analysis and amplification results and specificity are employed, and simultaneously decuple progressive dilution is used to detect the sensibility of the one-step method with multiple RT-PCR. The detecting method provided by the invention accomplishes reverse transcription and PCR reaction and simultaneously detects pathogen of two diseases by extracting the total nucleic acid of sugarcane, which has the advantages of simple operation, economy of time, low cost, high specificity and sensibility and the like.

Description

technical field [0001] The invention relates to a detection method for sugarcane diseases, mainly for sugarcane mosaic virus (Sugarcane mosaic virus, SCMV) and sugarcane ratoon dwarf disease (RatoonStunting Disease, RSD) pathogenic bacteria Leifsonia xylem Subsp.Xyli, Lxx) simultaneous detection method. Background technique [0002] Sugarcane mosaic virus (SCMV) and sugarcane ratoon dwarf disease (Ratoon Stunting Disease, RSD) are two major diseases that occur widely in the world. They are also quite common and serious in various sugarcane areas in my country. All Sugarcane cultivars are all affected to varying degrees. [0003] Sugarcane mosaic disease is caused by sugarcane mosaic virus. Sugarcane mosaic virus is a positive single-stranded RNA virus and one of the important members of Potavirus genus Potaviridae. There are many virus strains and large variations. It mainly harms sugarcane leaves. The development of chloroplast granules in infected plant cells is inhibited...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C07H21/00
Inventor 杨本鹏张显勇张树珍蔡文伟
Owner 广西康田农业科技股份有限公司
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