Multiple PCR detection kit for three bacterial seed-borne diseases and special primer for kit as well as multiple PCR detection method

A bacterial leaf spot and bacterial technology, which is applied in biochemical equipment and methods, microorganism-based methods, and microbial determination/inspection, etc. 3. Detection has limitations and other problems, to achieve the effect of saving detection cost and time, saving detection time and reducing detection cost

Active Publication Date: 2015-06-24
YUNNAN AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In 2000, bacterial fruit spot disease of thick-skinned muskmelons occurred on a large scale in Bayannaoer League, Inner Mongolia, with an average yield reduction of 46%, and the commercial melon rate was only 1/3; in the winter of 2002, melon bacterial fruit spot disease The seedling destruction rate that causes is also up to 30%~80%, causes serious econ

Method used

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  • Multiple PCR detection kit for three bacterial seed-borne diseases and special primer for kit as well as multiple PCR detection method
  • Multiple PCR detection kit for three bacterial seed-borne diseases and special primer for kit as well as multiple PCR detection method
  • Multiple PCR detection kit for three bacterial seed-borne diseases and special primer for kit as well as multiple PCR detection method

Examples

Experimental program
Comparison scheme
Effect test

Example Embodiment

[0080] Embodiment 1 The design of the special primer Xj for the rapid detection of tomato canker of the present invention and the detection of tomato canker by conventional molecular biological methods using the pair of special primers Xj.

[0081] (1) Design and synthesis of special primer Xj for detection of tomato canker

[0082] The special primer Xj for the detection of tomato canker is based on the 16SrDNA gene fragment in the chromosomal genome of tomato canker. DNAMAN6.0 is used for homology analysis to determine the highly conserved region, and then Primer 6.0 primer design software is used to design specific primers, The number is dedicated primer Xj, which is composed of the upstream primer of the dedicated primer Xj and the downstream primer of the dedicated primer Xj, the base sequence of the upstream primer of the dedicated primer Xj is shown in SEQ ID NO: 1, and the downstream primer of the dedicated primer Xj The base sequence of the primer is shown in SEQ ID N...

Example Embodiment

[0100] Example 2 Design of the special primer Dc for rapid detection of tomato bacterial leaf spot according to the present invention, and the detection of tomato bacterial leaf spot by conventional molecular biology method using the pair of special primers Dc.

[0101] (1) Design and synthesis of special primer Dc for detection of tomato bacterial leaf spot

[0102] Special primer Dc for detection of tomato bacterial leaf spot The special primer Dc is based on the 23S rDNA gene fragment in the chromosome genome of tomato bacterial leaf spot. DNAMAN6.0 is used for homology analysis to determine the highly conserved region, and then Primer 6.0 primers are used. Design software, design specific primers and number them as special primer Dc, which is composed of special primer Dc upstream primer and special primer Dc downstream primer, the base sequence of the special primer Dc upstream primer is as SEQ ID NO: 3 As shown, the base sequence of the downstream primer of the special pri...

Example Embodiment

[0106] Embodiment 3 is the design of the special primer At for the rapid and accurate detection of cucurbit bacterial fruit spot disease of the present invention and the detection of cucurbit bacterial fruit leaf spot by conventional molecular biology method with this special primer At.

[0107] (1) Special primer At and its design and synthesis

[0108] The special primer At for the detection of cucurbit bacterial fruit spot is based on the hrpB2 gene fragment in the chromosomal genome of cucurbit bacterial fruit spot, and DNAMAN6.0 is used to conduct homology analysis to determine the highly conserved region, and then use Primer 6.0 primer design software , design a specific primer, and number it as a special primer At, which is composed of a special primer At upstream primer and a special primer At downstream primer, the base sequence of the special primer At upstream primer is shown in SEQ ID NO: 5 , the base sequence of the downstream primer of the special primer At is sh...

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Abstract

The invention discloses a multiple PCR detection kit for three bacterial seed-borne diseases and a special primer for the kit as well as a multiple PCR detection method. The invention provides special primers aiming at the canker pathogens of tomato, the bacterial leaf spot pathogens of the tomato and the bacterial fruit spot pathogens of melons; the target product fragment of the special primer for detecting the canker pathogens of the tomato is 1086bp; the target product fragment of the special primer for detecting the bacterial leaf spot pathogens of the tomato is 491bp; and the target product fragment of the special primer AT for detecting the bacterial fruit spot pathogens of the melons is 290bp. The invention also provides a multiple PCR optimal reaction system for the three pathogens and reaction conditions. The multiple PCR detection kit is capable of quickly and simultaneously detecting the canker pathogens of the tomato, the bacterial fruit spot pathogens of melons and the bacterial leaf spot pathogens of the tomato without cloning, sequencing and sequence alignment, and is high in specificity, quick and accurate; and as a result, the detection cost and the detection time are saved, and the limitation of the single reaction is solved.

Description

technical field [0001] The present invention relates to the technical field of molecular biology detection of three kinds of plant pathogens, in particular to Clavibacter michiganensis subsp.michiganensis, tomato bacterial leaf spot (pseudomonas syringae pv.tomato) and melon bacterial fruit spot ( Acidovorax citrulli) multiplex molecular biology detection method. Background technique [0002] my country is the fourth largest importer and fifth largest exporter of agricultural products. However, with the further expansion of foreign trade, the transportation of plants and their products has become increasingly frequent, especially the most dangerous seeds. This has become a very important and urgent problem that endangers my country's biodiversity, ecological environment and national economy. The number and variety of alien invasive organisms are large. Already 529 species of alien organisms have invaded 34 provincial-level administrative regions in my country, leading to spe...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12Q1/04C12N15/11C12R1/01
CPCC12Q1/686C12Q1/689C12Q2600/16C12Q2537/143C12Q2565/125
Inventor 刘雅婷王金龙岳凯李小林谷安宇
Owner YUNNAN AGRICULTURAL UNIVERSITY
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