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Loop-mediated isothermal amplification detection primer set, method and kit for detection of Xanthomonas oryzae

A technology of Xanthomonas oryzae, ring-mediated isothermal, applied in the field of microbial detection, to achieve the effect of good specificity, rapid detection method and high sensitivity

Active Publication Date: 2019-03-19
HAINAN UNIVERSITY
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

At present, there is no specific, efficient and simple method for rapid detection of pathogenic bacteria in the field

Method used

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  • Loop-mediated isothermal amplification detection primer set, method and kit for detection of Xanthomonas oryzae
  • Loop-mediated isothermal amplification detection primer set, method and kit for detection of Xanthomonas oryzae
  • Loop-mediated isothermal amplification detection primer set, method and kit for detection of Xanthomonas oryzae

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Embodiment 1

[0032] 1 Materials and methods

[0033] 1.1 Materials:

[0034] Diseased rice seeds and leaves were collected from Luoniushan Farm, Haikou City, Hainan Province. Xanthomonas oryzae and Bacterial spot bacteria were preserved in the laboratory. Chemical reagents were purchased from Guangzhou Chemical Reagent Company and Sigma-Aldrich. DNA polymerase and amplification Reagents for amplification were purchased from New England Biolabs. The primers are listed in Table 1 and were synthesized by Sangon Bioengineering Co., Ltd.

[0035] Table 1. P1 primers used in this study

[0036]

[0037] 1.2 Equipment:

[0038] 1.5mL centrifuge tube, 0.2mL PCR tube, desktop room temperature centrifuge, vortex shaker, pipette, tip, electrophoresis equipment.

[0039] 1.3 Method:

[0040] 1.3.1 Rice total genome extraction

[0041] Extraction buffer (Buffer DL): 50mM Tris-CL (pH7.8), 0.35mM sorbitol, 5mM EDTA, 1% mercaptoethanol, 1% CTAB, 5mM NaCl.

[0042] 1a. Rice seedlings: cut 2-3cm lea...

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Abstract

The invention discloses a loop-mediated isothermal amplification detection primer set, method and kit for detection of Xanthomonas oryzae. Based on genome specific molecular marker characteristics ofXanthomonas oryzae, conservative specific DNA fragment xcp of Xanthomonas oryzae is acquired by means of a comparative genomics method; diseased rice seedlings and seeds are successfully detected through loop-mediated isothermal amplification; the method herein is quick, simple and efficient. The whole detection process has no need for special instruments and can be finished within 45-90 min in afield or lab. The method is high in sensitivity, suitable for detecting rice materials containing down to 10 bacteria, good in specificity, and is free of influence upon results due to other pathogensor plant genomes. The method is good in safety and free of harm and employs no organic agents or nucleic acid binding dyes; all agents are directly releasable to the environment; the method is suitable for detecting all possible pathogen propagating sources.

Description

Technical field: [0001] The invention belongs to the field of microorganism detection, and in particular relates to a detection primer set, a detection method and a detection kit for a ring-mediated isothermal amplification detection of Xanthomonas oryzae. Background technique: [0002] Rice (Oryza sativa) is the main food crop for more than half of the world's population, most of whom live in developing countries (Wei Shuya et al., 2013). China is the most populous country in the world, and rice is widely planted in China with the largest output. The yield of rice has a great impact on people's life and plays an important role in the development of social economy. It is understood. In my country, more than 60% of people use rice as their main food source, which fully demonstrates the important role of rice in people's lives. In my country, the south is the main planting place of rice, the "granary" that most people in our country rely on for survival, and it is also the ...

Claims

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/6844C12Q1/04C12N15/11
CPCC12Q1/6844C12Q1/689C12Q2531/119C12Q2563/107
Inventor 陶均陈银华何朝族郑琳琳李春霞牛晓磊冯世鹏
Owner HAINAN UNIVERSITY
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