Bacterium for inhibiting pathogenic bacteria of tobacco diseases and application
A technology of bacteria and tobacco, applied in the direction of application, bacteria, chemicals for biological control, etc., to achieve the effects of simple production process, good colonization effect, and wide sources
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Embodiment 1
[0131] - Acquisition of Bacillus velezensis WY2
[0132] A. Separation:
[0133] Take 0.4g of tobacco plant stem tissue that has been thoroughly sterilized on the surface, put it into a 2mL sterilized centrifuge tube together with sterilized steel balls, add 600μL of sterile water, and grind it with a tissue grinder for 3min at a frequency of 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;
[0134] B. Screening:
[0135] Using the plate confrontation method, use a hole puncher with a d...
Embodiment 2
[0144] ——Identification and preservation of Bacillus velezensis WY2
[0145] (1) Identification of Bacillus velezensis WY2
[0146]For the strain WY2 selected and bred 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 Sangon Biotechnology Co., Ltd. for sequence determination.
[0147] The test results are recorded as follows:
[0148] 1. Culture characteristics: The strain was cultured on NA plate medium at 28°C. In the ...
Embodiment 3
[0157] ——The control experiment of Bacillus velezensis WY2 on tobacco Fusarium root rot
[0158] The experiment was carried out in Chengjiang County, Yuxi City, Yunnan Province, and the tobacco variety was "Yunyan 87".
[0159] experiment method:
[0160] a. Fermentation: Activate the strain WY2 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 48 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 temperature incubator at 28°C for 72 hours , ...
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