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Bacillus velezensis Z002 as well as acquisition method and application thereof

A technology of Bacillus Velez, Z002, which is applied in the field of microbial application to achieve the effects of low cost, convenient application and good stress resistance

Pending Publication Date: 2021-02-05
YUNNAN ACAD OF TOBACCO AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, there are few reports on microorganisms used to prevent and treat mildew in the curing period, only Bacillus mucilaginosus (CN110384247A) and combined microbial agent (CN110250209A) are used as biological control products to control tobacco mildew in the curing period. to report

Method used

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  • Bacillus velezensis Z002 as well as acquisition method and application thereof
  • Bacillus velezensis Z002 as well as acquisition method and application thereof
  • Bacillus velezensis Z002 as well as acquisition method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] - Acquisition of Bacillus velezensis Z002

[0091] A. Separation: Take 0.4g of the stem tissue of tobacco plants that has been thoroughly sterilized on the surface after inspection, put them together with sterilized steel balls into a 2mL sterilized centrifuge tube, add 600μL of sterile water, grind for 3min with a tissue grinder, and the frequency is 30r / s. Gradually dilute the ground tissue fluid to 10 -4 , 200 μL of homogenate was applied to the separation medium (5.0 g of peptone, 3.0 g of beef extract, 8.0 g of sodium chloride, 15.0 g of agar, pH 7.0) of each gradient, and each treatment was repeated 4 times. After culturing in the dark at 28°C for 48 hours, pick a single colony according to the colony shape, color, etc., re-stretch and purify on the separation medium, and transfer the single colony cultured by the streak to the slant of the separation medium test tube for storage;

[0092] B. Screening: adopt the flat plate confrontation method, stick the mycel...

Embodiment 2

[0098] ——Identification and preservation of Bacillus velezensis Z002

[0099] (1) Identification of Bacillus velezensis Z002

[0100] For the strain Z002 selected above, biological analysis and molecular biological identification were carried out by conventional methods. The molecular identification method is as follows: Bacteria Genomic DNA Extraction Kit, see the kit instruction manual for the method. Primer F27 / R1492 was selected for PCR amplification, amplified under conventional conditions, the amplified product was recovered by gel, connected to the vector pEAZY-T5 Zero vector, heat-shock transformed Escherichia coli competent cell DH5α, and colonies were picked with M13F / M13R as primers Perform colony PCR identification. Positive clones were sent to Shanghai Yingjun Biotechnology Co., Ltd. for sequence determination.

[0101] The test results are recorded as follows:

[0102] 1. Culture characteristics: The strain was cultured on NA plate medium at 28°C. In the earl...

Embodiment 3

[0111] ——The control test of Bacillus velezensis Z002 on tobacco mildew during the curing period was carried out in Fengyi Town, Xiaguan City, Dali Prefecture, Yunnan Province. The tobacco variety was "Honghua Dajinyuan", and the tobacco leaves were middle blade.

[0112] experiment method:

[0113] a. Fermentation: Activate the strain Z002 from a -80°C ultra-low temperature refrigerator in a slant medium (10.0g peptone, 3.0g beef extract, 5.0g sodium chloride, 15.0g agar, pH 7.0), and culture in a constant temperature incubator at 28°C 48h; Pick a ring of bacteria and inoculate it into a 50mL centrifuge tube containing 10mL of seed culture solution (10.0g of peptone, 3.0g of beef extract, 5.0g of sodium chloride, 15.0g of agar, pH 7.0), and place in a constant temperature incubator at 28°C Cultivate for 24 hours; then transfer the seed solution to 1000mL fermentation medium (peptone 10.0g, beef extract 3.0g, sodium chloride 5.0g, agar 15.0g, pH 7.0), and place in a constant ...

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Abstract

The invention discloses bacillus velezensis Z002 as well as an acquisition method and application thereof. The bacillus velezensis Z002 is collected in China Center for Type Culture Collection (CCTCC), and the collection number is CCTCC No: M 2020567. The acquisition method comprises separation, screening and purification processes. Endophytic bacteria are firstly separated from tobacco plant tissue and subjected to dark culture, the endophytic bacteria antagonizing the tobacco mildew pathogen rhizopus oryzae are screened out, and finally purification is carried out to obtain the bacillus velezensis Z002. The application of the bacillus velezensis Z002 comprises fermentation, inoculation and modulation processes. Firstly, strains are cultured to obtain a fermentation inoculant, the fermentation inoculant is sprayed to tobacco petioles through a sprayer, tobacco leaves are put into a curing barn, and the tobacco leaves are cured according to a normal curing process. The strain bacillusvelezensis Z002 can effectively control tobacco mildew in the curing period, and the relative control effect is 80% or above. Secondly, the bacillus velezensis Z002 also has very good bacteriostatic effects on Ralstonia pseudosolanacearum, Phylophthora nicotianae, Botrytis cinerea, Sclerotinia sclerotiorum, Thanatephorus cucumeris, Fusarium oxysporum, Alternaria alternata and the like. The bacillus velezensis Z002 has good control effect and higher application and popularization value.

Description

technical field [0001] The invention belongs to the technical field of microbial application, and relates to a strain of Bacillus Velez Z002, an acquisition method and its application, in particular to a strain of Bacillus Velez Z002 capable of efficiently preventing and curing mildew in the baking period, and an acquisition method thereof And its application in the prevention and treatment of tobacco mildew in barn. Background technique [0002] Tobacco mold disease in the curing period is a fungal disease caused by Rhizopus oryzae infection, which can damage tobacco leaf petiole and mesophyll tissue, mainly leaf base mildew type, with an incidence rate of more than 30% and a loss of 20%. %about. Therefore, the harm of tobacco mildew to tobacco leaf production is very serious. [0003] The control methods for tobacco mildew during the curing period include chemical agents, improvement of curing equipment and biological control. [0004] Although the chemical control meth...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N1/02A01N63/22A01P3/00C12R1/07
CPCC12N1/20C12N1/02A01N63/22C12R2001/07C12N1/205
Inventor 卢灿华夏振远马俊红晋艳盖晓彤雷丽萍陈颐苏家恩鲁子贤姜宁蔺忠龙
Owner YUNNAN ACAD OF TOBACCO AGRI SCI
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