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Method for direct rapid detection and identification of banana-root nematode in soil or other media

A technology for banana perforating nematodes and soil, which is applied in the field of molecular biology detection and identification, and achieves the effects of high detection sensitivity and good purification effect.

Inactive Publication Date: 2012-10-03
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there are few reports on direct detection and identification of nematodes in soil by molecular biological methods

Method used

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  • Method for direct rapid detection and identification of banana-root nematode in soil or other media
  • Method for direct rapid detection and identification of banana-root nematode in soil or other media
  • Method for direct rapid detection and identification of banana-root nematode in soil or other media

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1 Rapid detection and identification of the banana borer nematode from soil samples containing the banana borer nematode

[0030] Test materials: Banana perforator nematodes were collected from the rhizosphere soil of Anthurium anthurium, cultured and preserved on carrot callus in a 25°C incubator in our laboratory; the soil was collected from the rhizosphere soil of bananas in the field.

[0031] The specific implementation steps are as follows:

[0032] 1. Extraction of total DNA from mixed samples of banana borer nematode and soil

[0033] 1.1 Weigh 0.5 g of the above-mentioned field soil sample, put it into a 15 mL sterilized conical centrifuge tube, and add an equal weight of small glass beads with a diameter of 0.5 mm;

[0034] 1.2 The banana perforator nematodes were isolated from the carrot callus group where the nematode was cultivated, and 50, 5, 4, 3, 2 and 1 banana perforator nematodes were picked under the microscope and put into different soils ...

Embodiment 2

[0055] Example 2 Rapid detection and identification of the banana borer nematode directly from the banana rhizosphere soil where the banana borer nematode disease occurred

[0056] Test materials: In this laboratory, the rhizosphere soil of Banana brasiliensis inoculated with the nematode and diseased in the greenhouse was potted.

[0057] The specific implementation steps are as follows:

[0058] Take about 500-1 000 g of the rhizosphere soil of the diseased Brazilian banana, mix it well, weigh 5 parts of 0.5 g soil (No. 1-5), and weigh 0.5 g of sterilized soil (No. In a sterilized centrifuge tube, add an equal weight of small glass beads with a diameter of 0.5 mm; other steps are the same as in Example 1, and the test results are shown in figure 2 . Depend on figure 2 It can be seen that the DNA of the brazilian rhizosphere soil infected with the banana nematode has the target fragment after PCR detection, while the soil DNA without the banana nematode does not detect...

Embodiment 3

[0059] Example 3 Rapid detection and identification of the banana borer nematode from samples mixed with the substrate

[0060] Experimental material: The preparation and acquisition of the banana perforator nematode were the same as in Example 1, and the substrate was purchased from Guangzhou Fangcun Lvyuan Horticultural Plant Material Farm.

[0061] The specific implementation steps are as follows:

[0062] Weigh 0.5 g of matrix, put it into a 15 mL sterilized centrifuge tube, and add an equal weight of small glass beads with a diameter of 0.5 mm. Other steps are with embodiment 1. See the test results image 3 .

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Abstract

The invention discloses a method for direct rapid detection and identification of banana-root nematode in soil or other media. The method includes extracting DNA (deoxyribonucleic acid) in a sample to be detected, using a Wizard DNA clean-up systems kit of Promega to purify the DNA, using the purified DNA as a template, using SEQs ID NO.1 and NO.2 as primer for PCR (polymerase chain reaction) amplification, and confirming presence of the banana-root nematode in the sample to be detected if an objective band 518bp is obtained by amplification. The method is applicable to detection of banana culture soil or other media, detection results are accurate, sensitivity is high, and the method is widely applicable and has broad application prospect.

Description

technical field [0001] The invention relates to the technical field of molecular biology detection and identification, in particular to a method for quickly detecting and identifying the banana perforator nematode directly from soil or other substrates. Background technique [0002] Banana perforator nematode ( Radopholus similis (Cobb) Thorne, 1949) is a migratory plant endoparasitic nematode, with more than 250 species of host plants reported (O’Bannon, 1977), which seriously damages banana ( Musa spp.), citrus ( Citrus spp.), pepper ( Piper nigrum ), ginger ( Zingier officinale ) and other economic plants and ornamental plants (Luc et al, 1990; Richardson et al, 1993; Uchida et al, 2003; Sundararaju et al, 1979), Banana perforator nematodes are widely distributed in most banana production areas in the tropics and subtropics In greenhouses in temperate and temperate regions, the distribution of this nematode has been reported in more than 90 countries and region...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 谢辉徐春玲刘一帆李静李冬丽赵传波
Owner SOUTH CHINA AGRI UNIV
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