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Co-culture method of trichoderma atroviride and bacillus subtilis and an application of co-metabolite obtained by method

A technology of Bacillus and Bacillus subtilis, applied in botany equipment and methods, biochemical equipment and methods, microorganism-based methods, etc., can solve problems such as the difficulty of creating new microbial pesticides

Pending Publication Date: 2020-03-31
浙江华智生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, at present, the combination of two types of microorganisms at home and abroad is mainly to ferment the two kinds of microorganisms separately to prepare bacterial agents, and then combine or mix them in proportion. Although it can improve the control effect and stability, it is difficult to create new microbial pesticides.

Method used

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  • Co-culture method of trichoderma atroviride and bacillus subtilis and an application of co-metabolite obtained by method
  • Co-culture method of trichoderma atroviride and bacillus subtilis and an application of co-metabolite obtained by method
  • Co-culture method of trichoderma atroviride and bacillus subtilis and an application of co-metabolite obtained by method

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Experimental program
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Effect test

Embodiment 1

[0042] The co-cultivation method of embodiment 1, Trichoderma and bacillus

[0043] (1) Preparation of plate strains:

[0044] Weigh 200g potatoes and cut into 1cm 2 The small pieces were boiled on low heat for 30 minutes, filtered through 4 layers of gauze, and the juice was reserved. Add 20 grams of glucose, 20 grams of agar powder, distilled water to 1 liter, and sterilize under high pressure at 121°C for 30 minutes to prepare PDA medium. Inoculate Trichoderma dark viridans SG2403 on a PDA medium plate, in an incubator at 28°C, and culture it upside down for 2-3 days.

[0045] Weigh 10 grams of peptone, 8 grams of yeast powder, 10 grams of NaCl, 20 grams of agar powder, distilled water to 1 liter, and sterilize at 121 ° C for 30 minutes to prepare LB solid medium, and inoculate Bacillus subtilis 22 in LB culture In the basal plate, in a 30°C incubator, culture upside down for 1-2 days.

[0046] (2) Preparation of liquid strains:

[0047] Scrape Trichoderma spores from...

Embodiment 2

[0052] Embodiment 2, co-cultivation fermented liquid antagonism activity assay of wheat scab

[0053] Trichoderma Bacillus co-cultivation fermentation liquid and single Trichoderma fermentation liquid, Bacillus fermentation liquid (referring to: the Trichoderma conidia suspension of 1 milliliter is added in the aforementioned co-cultivation fermentation medium, 28 ℃, 180 rpm, cultivated for 5 days, to obtain a single Trichoderma fermentation broth; add 1 ml of Bacillus bacteria liquid to the aforementioned co-cultivation fermentation medium, 28 ° C, 180 rpm, and cultivate for 5 days to obtain a single Bacillus fermented liquid.) With 0.22 micron microporous membrane filtration sterilization, join the PDA medium (the formula of this medium is the PDA cultivation mentioned in the preparation of front plate strains) with the amount of 200 microliters / 10 milliliters In the base formula), the blank PDA was used as a control, and the center of the plate was inoculated with Fusariu...

Embodiment 3

[0056] Embodiment 3, the response surface optimization of co-cultivation formula

[0057] (1) Plackett-Burman design (PBD): Based on the initial fermentation medium (20 g corn flour / liter, 20 g yeast powder / liter, 20 g molasses / liter), use the Plackett-Burman design to determine the effect on fermentation The main factor of liquid antibacterial rate. A total of 12 experiments were run with 7 actual variables (X1, X2, X3, X6, X7, X8 and X11) and 4 dummy variables (X4, X5, X9 and X10) (Table 1). In this study, the actual variables are set to two levels, which are rotational speed (rpm) (X1) (180, 200), temperature (°C) (X2) (28, 30), initial pH value (X3) ( 6.5, 7.5), corn flour concentration (g / L) (X6) (10, 20), molasses concentration (g / L) (X7) (10, 20), yeast powder concentration (g / L) (X8) ( 10, 20) and KCl concentration (g / L) (X11) (0,1) (Table 1). The inhibitory rate of fermentation broth to wheat head blight was taken as the response value.

[0058] Table 1 Plackett...

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Abstract

The invention discloses a co-culture method of trichoderma atroviride and bacillus subtilis and an application of a co-metabolite obtained by the method. According to the method, the trichoderma atroviride SG3403 and bacillus subtilis 22 strains are used as co-culture objects, the inhibition rate of a co-culture fermentation liquid against fusarium graminearum is used as a response value, and thefermentation conditions such as culture temperature, a rotation speed, a pH, a corn flour content, a molasses content, a yeast powder content and a KCl content are optimized by a response surface method to obtain the following best fermentation conditions: the culture temperature is 28 DEG C, the rotation speed is 214 rpm, the pH value is 6.8, the yeast powder content is 10 g / L, and the molasses content is 27.6 g / L. The co-culture fermentation liquid cultivated by the formula provided by the invention improves the resistance to the fusarium graminearum, a substance not found in a single culture is found in the co-culture fermentation liquid, a seed coating agent prepared from the substance promotes the growth of wheat, and a novel option is provided for the development of biological pesticides and biological seed coating agents.

Description

technical field [0001] The present invention relates to a co-cultivation method of Trichoderma and Bacillus and the use of co-metabolites thereof; in particular to an optimization method of co-cultivation technology of Trichoderma and Bacillus for the prevention and treatment of wheat scab and the promotion of wheat growth and its metabolism The antagonism of the product to wheat head blight and the application in promoting the growth of wheat seedlings. Background technique [0002] Trichoderma and Bacillus are important agricultural and industrial microorganisms, especially as biocontrol agents and plant growth promoters. However, the single use of the two microorganisms has certain limitations. Trichoderma mainly controls soil-borne diseases through competition, hyperparasitism and induction of resistance, while Bacillus mainly controls leaf diseases through antibiosis. From the secondary metabolic spectrum, the two are also obviously different. Trichoderma can produce...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N1/20C12N1/14A01N63/38A01N63/22A01P3/00A01P21/00C12R1/125C12R1/885
CPCC12N1/14C12N1/20
Inventor 李婷婷陈捷唐家全
Owner 浙江华智生物科技有限公司
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