Preparation method and application of entomogenous fungus antibacterial peptide
A technology of insect-borne fungi and antibacterial peptides is applied in the preparation methods of peptides, botanical equipment and methods, applications, etc., to achieve the effects of short production cycle, low production cost, and high food safety.
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Embodiment 1
[0035] Example 1 Effects of Different Fermentation Mediums on the Production of Active Peptides by the Entomogenic Fungus Cordyceps cicadae
[0036] (1) Entomophagous fungal strains: Cordyceps cicadae ( O. sobolifera ) 022007-9 strain;
[0037] (2) Strain activation and seed preparation: Pick mature spores of the preserved strains and culture them on a PSA slope, and activate them at 25±1°C for 7-10 days. Add sterilized distilled water to the slope, wash the spores down, shake vigorously for 3 to 5 minutes to make a spore suspension (seed), and calculate the spore concentration by blood cell counting method for later use;
[0038] (3) Fermentation culture: 6×10 7 Inoculate 1ml of the spore suspension into the following three different 250mL fermentation broths, shaker speed 120rpm / min, culture at 25±1°C for 7 days, three repetitions each, shaker speed 120rpm / min, 25±1°C Culture at 1°C for 7 days; 3 replicates each; selected fermentation medium: ① 200g potato, 20g sucrose...
Embodiment 2
[0044] Example 2 Preparation steps of entomogenic fungal antimicrobial peptide products
[0045] The preparation steps of the entomogenic fungal antimicrobial peptide product are as follows:
[0046] (1) Entomophagous fungal strains: Cordyceps cicadae ( O. sobolifera ) 022007-9 strain;
[0047] (2) Strain activation and seed preparation: Pick mature spores of the preserved strains and culture them on a PSA slope, and activate them at 25±1°C for 7-10 days. Add sterilized distilled water to the slope, wash the spores down, shake vigorously for 3 to 5 minutes to make a spore suspension (seed), and calculate the spore concentration by blood cell counting method for later use;
[0048] (3) Fermentation culture: 6×10 7 1 ml of spores were inoculated into 250 ml of fermentation broth, the shaking table was rotated at 120 rpm / min, and cultured at 25±1°C for 7 days;
[0049] The fermentation medium used in the step (3) is: peptone 5g, sucrose 20g, KH 2 PO 4 1g, MgSO 4...
Embodiment 3
[0056] Example 3 Tricine-SDS-PAGE electrophoresis detection of samples
[0057] The purity and molecular weight estimation of purified samples were carried out with reference to the Tricine-SDS-PAGE electrophoresis method of Sch?gger et al. (1987) [Sch?gger H, von Jagow G. Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis for the separation of proteins in range from 1 to 100 kDa. Analytical Biochemistry, 1987, 166: 368-379]. 4% stacking gel, 16.5% separating gel. Mix the sample and sample buffer at 1:1 (volume ratio), heat in a boiling water bath for 5 min, and centrifuge at 5,000 × g for 7 min.
[0058] Load 15 μL of supernatant into each well. Electrophoresis at a voltage of 30 volts until the sample completely enters the separation gel from the stacking gel, then the voltage is increased to 80 volts for electrophoresis, and the electrophoresis is stopped when the bromophenol blue indicator migrates to the bottom of the separation gel.
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