Paenibacillus polymyxa microbial inoculum as well as preparation method and application thereof
A technology of Paenibacillus polymyxa and Bacillus, which is applied in the field of microorganisms, can solve the problems of enhanced drug resistance of pathogenic bacteria, influence of beneficial microorganisms, and changes in soil physical and chemical properties, and achieve significant inhibitory effects, reduce incidence, and inhibit growth and development.
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[0037] On the other hand, the embodiment of the present invention also provides a preparation method of the above bacterial agent, the method includes the following steps, under aseptic conditions, pick a single colony of Paenibacillus polymyxa Cs12 and insert it into a sterilized In NA or PDA medium, cultivate in dark at 28-37°C, 75% RH for 24-48h to obtain purified colonies; under sterile conditions, pick purified colonies and inoculate them in sterilized NB or PDB medium. Shake culture at 28°C and 180rpm for 24 hours to obtain seed bacteria; inoculate the seed bacteria in NB or PDB medium, and ferment at 28°C and 180rpm for 48 hours to form a fermentation broth to obtain the bacterial agent. Wherein Paenibacillus polymyxa Cs12 is first cultured in NA medium for a period of time to obtain a single colony of purified Paenibacillus polymyxa Cs12, which is inoculated in NB or PDB medium by picking the single colony to obtain Seed fungus culture medium for subsequent fermentatio...
Embodiment 1
[0043] Screening, isolation, purification and identification of Paenibacillus polymyxa Cs12:
[0044] (1) On August 15, 2020, healthy saffron rhizosphere soil was collected from a grape-saffron intercropping field in the South Taihu Modern Agriculture Demonstration Park in Huzhou, Zhejiang. Weigh 0.5g of soil and add 2mL of sterile water, shake at 180rpm / min for 20min to obtain a suspension, and use the RoToR HDA Colony High-throughput Screening Workstation (SINGER, UK) to automatically spot samples on a 96-well plate, with Fusarium oxysporum in the middle , four soil suspensions were placed equidistantly in a square around them. After the sampling was completed, the plate was sealed with a parafilm, and cultured in the dark at 37°C for 2-3 days, and the colonies with obvious antibacterial zones were selected for isolation and purification.
[0045] (2) Pick colonies with obvious inhibition zone, inoculate them on NA medium by streaking method, and purify them. Pick the purif...
Embodiment 2
[0055]Disease resistance experiment of the bacterial agent of Paenibacillus polymyxa Cs12:
[0056] (1) Take the strain of Fusarium oxysporum preserved on a slant, and under aseptic conditions, use an inoculation needle to pick a little mycelium and inoculate it on PDA medium for activation, and cultivate it in a dark environment at 26±2°C for 5-7d; A sterilized hole puncher was used to make bacterial blocks, which were inoculated into PDB liquid medium for culture, and 1 bacterial block was inoculated per 10ml of PDB. Incubate at 30°C, 180rpm / min for 4-5d. The cultured Fusarium oxysporum fermentation broth was filtered with two layers of sterile gauze in an ultra-clean bench. Draw 100 μL of spore suspension, count the number of spores with a hemocytometer, and adjust to F.oxysporum 10 5 -10 6 spores / mL suspension.
[0057] (2) Cs12 strains cryopreserved in 25% glycerol, under aseptic conditions, draw 100 μL and spread it on NA medium, culture in dark at 28-37°C and 75% hu...
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