Endophytic fungus paraconiothyrium brasiliense strain and application thereof
A technology of endophytic fungi and strains, applied in the direction of fungi, microorganisms, microorganisms, etc., to achieve the effect of optimizing culture conditions
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Embodiment 1
[0023] Isolation and preservation of Paraconiothyrium brasiliense:
[0024] In October, 2011, the insects of S. sinensis were collected in Dajiuhu National Wetland Park in Shennongjia, Hubei Province. The insects were rinsed with distilled water, and sterilized in 75% ethanol for 1 minute under aseptic conditions, and then rinsed with sterile water. Rinse 4 times, then sterilize with 3% sodium hypochlorite for 1 min, rinse with sterile water 4 times, then use sterile scissors to cut the worm body from the abdomen, take the intestine, cut it, use the inoculation loop dipped in its intestinal fluid, and streak onto the PDA medium plate Place it on a constant temperature culture at 28°C and observe regularly. After the mycelium grows, use the method of purifying the top of the mycelium to pick the top of the mycelium and purify it on a new PDA medium plate. When colonies of different colors or shapes grow After coming out, continue to carry out purification operation with mycelia...
Embodiment 2
[0026] Identification of Paraconiothyrium brasiliense:
[0027] Sequence determination of Cyclospora brasiliensis strains: the strains were cultured in PDA liquid and filtered to collect mycelium, and an appropriate amount of mycelium was quickly ground into powder with liquid nitrogen, and then the Ezup column genomic DNA extraction kit (fungi) was used to extract the strain DNA, the extracted DNA was eluted with sterile double distilled water for PCR amplification. A pair of universal primers ITS5 / ITS4 was used for PCR amplification of ITS rDNA.
[0028] Forward primer ITS5: 5'-GGAAGTAAAAGTCGTAACAAGG-3', reverse primer ITS4: 5'-TCCTCCGCTTATTGATATGC-3'.
[0029] Use the ready-to-use PCR amplification kit to amplify. The PCR reaction system is: 1 μL of template DNA, 2 μL of dNTP (2.5 mmol / L each), 1 μL of each primer, 2 μL of 10×PCR buffer, 0.2 μL of Taq enzyme, ddH 2 O 12.8 μL, total system 20 μL. The reaction program was: denaturation at 94°C for 1 min, annealing at 56°C ...
Embodiment 3
[0032] A method for preparing chuanxiongdole from the metabolite of Cyclospora brasiliensis, comprising the following steps:
[0033] Step 1: Cultivation and fermentation of Microsporum brasiliensis: Inoculate the bacterial strain on a PDA medium plate, cultivate it in a constant temperature incubator at 28°C for 2 days, and pick a piece of bacterial body about 1cm×1cm with an inoculation needle to inoculate it on the In a Erlenmeyer flask (500 mL) equipped with 200 mL of PDA liquid medium, at a temperature of 28° C. and a rotation speed of 180 rpm, the seeds were cultured on a constant temperature shaking shaker for 5 days to obtain a seed solution. The seed solution was inoculated into a conical flask (500 mL) equipped with 200 mL of PDA liquid medium according to the inoculum amount of 5 mL, and continued to cultivate for 20 days with a constant temperature shaking shaker at a temperature of 28 ° C and a rotation speed of 180 rpm.
[0034] Step 2: Extraction of secondary me...
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