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A method for species-level classification of pear black spot fungus using multiple marker genes

A pear black spot fungus and marker gene technology, applied in the field of bioengineering, can solve the problems of non-sporulation and difficulty in species identification

Active Publication Date: 2019-08-06
INST OF FRUIT & TEA HUBEI ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In the process of species-level identification, it was found that the conidia of the microspore species Alternaria had polymorphism, and the morphological characteristics were easily affected by culture conditions such as temperature, humidity, and light; in addition, the conidia from the host and from the medium There are large differences in morphology; the sporulation phenotype is relatively easy to observe, but in the actual operation process, there are some transitional behaviors; there is also a class of Alternaria strains with little or no sporulation, all of which are given bands for species identification. it must be difficult

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  • A method for species-level classification of pear black spot fungus using multiple marker genes
  • A method for species-level classification of pear black spot fungus using multiple marker genes
  • A method for species-level classification of pear black spot fungus using multiple marker genes

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Embodiment Construction

[0016] In order to make the technical means, creative features, objectives and effects achieved by the present invention easy to understand, the present invention will be further explained below with reference to the drawings and specific examples.

[0017] 1. Isolation and cultivation of strains

[0018] Collect leaves and fruits with typical black spot symptoms in the field, and isolate pathogenic bacteria by tissue separation method. Take about 0.5cm at the junction of diseased spot and healthy tissue 2 Large and small tissues, first sterilize in 0.3% sodium hypochlorite solution for 30s, then 75% alcohol for 30s, then rinse 3 times in clean water, absorb water on sterile filter paper, place on PDA plate, culture at 25℃ under normal light Cultivate in a box.

[0019] 2. Isolation of monospores (single hyphae) of strains

[0020] After the colony grows on the PDA plate for 5 days, rinse the colony repeatedly with 1ml of sterile water, dilute the spore solution to a certain concentr...

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Abstract

The invention discloses a method for species level classification of alternaria kikuchi-aria by means of multiple marker genes. The method comprises the steps of 1, conducting PCR amplification on a strain to be detected by means of specific primers of the marker genes LEU / GPD and HIS; 2, conducting sequencing after an amplification product is connected with PUC19-vector; 3, conducting system analysis on the LEU / GPD+HIS comprehensive sequence by means of MEGA software, so that different alternaria kikuchi-aria strains are classified. By the adoption of the method, species level classification of alternaria kikuchi-aria can be achieved accurately and quickly.

Description

Technical field [0001] The invention belongs to the technical field of bioengineering, and specifically relates to a method for classifying Pear black spot disease bacteria by using multiple marker genes. Background technique [0002] Pear black spot is caused by Alternaria alternata fungus infection, which mainly damages fruits, leaves and new shoots. The disease occurs widely worldwide. Especially in Asia, Japan, South Korea, and China are seriously affected. [0003] In 1933, Tanaka first reported pear black spot in Japan and named the pathogen Alternariakikuchiana Tanaka. As early as 1920, Nagano reported the scientific name of the bacteria Alternaria gaisenNagano, therefore, according to the nomenclature, simmon believes that A.kikuchiana is a late synonym of A.gaisen. [0004] There are currently 9 species of Alternaria isolated from pear fruits during storage in the world, and 5 species reported in China, namely Alternaria gaisen, Alternaria alternata, Alternaria tenuissima...

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6895C12Q1/04C12R1/645
CPCC12Q1/6895
Inventor 朱红艳秦仲麒张晓旭伍涛李先明涂俊凡杨夫臣刘政朱婷
Owner INST OF FRUIT & TEA HUBEI ACAD OF AGRI SCI