Method for controlling pathogen of candidatus liberibacter in seedlings through composite microorganism treatment and application
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A compound microbial inoculum and compound microorganism technology are applied in the field of citrus virus-free seedling breeding to achieve the effects of improving the detoxification efficiency and solving the bottleneck problem.
Active Publication Date: 2020-10-30
HUAZHONG AGRI UNIV
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[0005] Since there is still no very effective control method for citrus huanglongbing, cutting down diseased trees, killing psyllids to control the vector and planting healthy and virus-free citrus seedlings are usually used for prevention an
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Embodiment 1
[0040] Example 1 strain identification
[0041] The microbial strains involved in this application were provided by the State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University. In this example, through 16S rDNA amplification, construction of expression vectors and sequencing, it was found that strain TD2-1 had a high degree of homogeneity with Bacillus Velez. Origin, TD2-2 has a high degree of homology with Bacillus amyloliquefaciens.
[0042] 1. Molecular biological identification of strains
[0043] 1) Extraction of bacterial genomic DNA
[0044] After the activation of the strain, take 2ml of the bacterial liquid and centrifuge to collect the bacterial cells, add 350μl of TE buffer solution containing 10mg / ml lysozyme to the pellet, mix gently, and incubate at 37°C for 30min. Then refer to the CTAB method to extract bacterial genomic DNA.
[0045] 2) Amplification of the 16S rDNA fragment of the strain
[0046] Using bacterial genomic DNA as ...
Embodiment 2
[0066] Example 2 Study on the inhibitory effect of compound bacterial agent treatment on the pathogenic source of citrus Huanglongbing
[0067] Preparation of microbial agent by mixed fermentation: pick single colonies of TD2-1 and TD2-2, respectively in LB medium, 200rpm, 37 ° C culture, the OD 600nm =1 Equal amount of TD2-1 and TD2-2 respectively get 10ml and add 20g brown sugar, add in autoclaved sterilized wheat bran (1KG)+rice bran (1KG)+corn starch (0.4KG)+soybean meal (0.2KG), After mixing according to the material-to-water ratio of 1:1, solid fermentation at 30°C for 7 days, during which, the culture was stirred once a day, and the culture was crushed. After drying at 60°C, the culture was crushed into fine powder with a pulverizer to detect live bacteria. The number is 20 billion / g, and it is ready for use. When used, dissolve in water according to the required dosage, 0.5L per plant.
[0068] 1. Bacteria treatment plan
[0069] Shatangju and Nanfeng tangerine seed...
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Abstract
The invention belongs to the technical field of breeding of citrus virus-free seedlings and particularly relates to a method for controlling a pathogen of candidatus liberibacter in seedlings throughcomposite microorganism treatment and an application. Citrus Huanglongbing is a destructive disease in citrus production all over the world, and the pathogen of the citrus Huanglongbing is caused by difficult-to-culture candidatus liberibacter. According to the method, scions are provided for cultivation of virus-free seedlings or non-toxic or low-toxic shoot tip materials are provided for micro-shoot grafting and detoxification of shoot tips by reducing the pathogen concentration or controlling the pathogen of the candidatus liberibacter in citrus seedlings, so that the detoxification efficiency can be greatly improved, and effective means are provided for solving the bottleneck problem of virus-free improved variety breeding of citrus. Through high-strength application of a bacillus complex, a method for controlling and reducing the pathogen concentration of the Huanglongbing is found; an important foundation is laid for subsequent micro-shoot grafting and detoxification work; the targets of reducing the pathogen concentration of a maternal plant and improving the detoxification effect are achieved; and safe and healthy shoot tip scions are quickly provided for industry, therebyachieving the disease-free improved variety of the citrus.
Description
technical field [0001] The invention belongs to the technical field of citrus virus-free seedling breeding, and in particular relates to a method and application of compound microorganism treatment and control of phloem bacillus pathogen in seedlings. Background technique [0002] Citrus Huanglongbing (HLB) has seriously hindered the development of the global citrus industry. Due to the difficulty in culturing its pathogenic bacteria in vitro, the research on the pathogenic mechanism of HLB has slowed down. At present, there is no targeted control method. [0003] Citrus Huanglongbing (HLB), also known as citrus huanglongbing (HLB), is a plant disease caused by bacteria of the genus Candidatus Liberibacter. At present, citrus huanglongbing has occurred and spread in more than 50 countries in Asia, Africa, Oceania, South America, and North America, and it is still expanding. In the United States and Brazil, Huanglongbing has had a devastating impact on the development of the...
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