Nematicidal compound derived from trichoderma virens as well as preparation method and application thereof
A compound, the technology of Trichoderma viridans, is applied in the fields of botany equipment and methods, biochemical equipment and methods, nematicides, etc. It can solve the problems of strong lethality of beneficial organisms, lack of ecological awareness, serious environmental pollution, etc., and achieve remarkable The effects of broad-spectrum nematicidal activity and good prospects for industrial promotion and application
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Embodiment 1
[0025] Example 1 Isolation and Identification of Trichoderma viridans (H.virens H22)
[0026] The applicant obtained a strain of Trichoderma viridans (Hypocrea virens H22, abbreviated as H.virens H22) which has a good poisonous effect on nematodes from the mangrove rhizosphere soil samples in Shenzhen Bay, Guangdong Province. The strain was released in September 2011. It was preserved on the 2nd at the China Center for Type Culture Collection of Wuchang Luojia Mountain, Wuhan City, Hubei Province, China, and the preservation number is CCTCCNO: M 2011303.
[0027] Sprinkle 1.0g soil sample on the culture medium (medium formula is CMA medium: 20g corn flour, 15g agar powder, 1000ml water), insert about 1000 second-instar larvae of M. Check and use a needle to pick up the single fungal conidia on and around the dead insect body and place them on a CMA plate containing 50 ppm of streptomycin sulfate, and purify to obtain the fungus.
[0028] The bacterial strain morphological cha...
Embodiment 2
[0051] Embodiment 2 The cultivation of Trichoderma viridans (H.virens H22)
[0052] (1) Culture by solid culture method
[0053] The medium formula is potato agar medium (PDA medium): 200g potato, 20g glucose, 20g agar powder, 1000mL water;
[0054] In each 90mm petri dish, pour about the thick PDA medium that accounts for the height of 1 / 3 of the petri dish, Trichoderma viridans (H.virens H22) mycelium is inoculated on the PDA medium, after sealing with parafilm Cultivate at 25-28°C for 2-3 days. Store the strains in a refrigerator at 4°C until use, or use them directly with spore culture solution.
[0055] (2) Culture by liquid culture method
[0056] The medium formula is CMA culture medium: corn flour 20g, distilled water 1000mL.
[0057] Each 500mL triangular flask was filled with 200mL of CMA culture solution, sterilized by damp heat at 121°C for 20min, and after cooling, 5 pieces of activated Trichoderma viridans discs were inserted into each bottle, and cultured in...
Embodiment 3
[0058] The preparation of embodiment 3 compound viridiol
[0059] Take the CMA culture solution (20g corn flour, 1000ml water) of Trichoderma virens (H.virens H22) and cultivate it in the dark at 180rmp 27.5°C for 7 days. Ethyl acetate was extracted three times, and the crude extract of biocontrol bacteria metabolites was obtained by rotary evaporation at 37°C.
[0060] Use 200-300 mesh silica gel column chromatography to fractionate the crude extract, first elute with low polarity chloroform, and then follow the volume ratio of methanol: chloroform as 1:9, 2:8, 3:7, 4 :6, 5:5, 6:4, 7:3, 8:2, 9:1 and methanol in order to increase the polarity of the eluent. Every 100ml collects a fraction, detects by TLC, merges the fractions with the same ratio shift value, and concentrates by rotary evaporation. When the solution of chloroform:methanol=7:3 is used as developing agent to carry out TLC detection, if the point with Rf value of about 0.3 (detection by ultraviolet light colorim...
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