Intestinal cellulose-degrading fungi of dragonfly larvae and their applications
A technology of cellulose and cellulase, applied in the biological field, can solve problems such as research on cellulase activity
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Embodiment 1
[0024] Embodiment 1, isolation and screening of bacterial strain
[0025] The tested yellow dragonfly larvae were collected from the river near the campus of Zhejiang Normal University. The surface of healthy yellow dragonfly larvae was sterilized with 75% (volume %) alcohol for 2 minutes, rinsed with sterile water for 3 times, and the intestinal tract obtained after dissection was put into a sterile mortar for grinding, and the grinding liquid was gradiently diluted with sterile water into 10 -1 , 10 -2 , 10 -3 , take 0.2mL of each gradient dilution solution and spread it on MEA medium (recipe: malt 20g, sucrose 20g, agar 20g, peptone 1g, distilled water to 1L, pH 7.0) plate, and culture in a 28°C incubator. After the colony grows, pick a small amount of hyphae from the edge of the colony, and transfer it to the MEA medium plate again, thus obtaining a pure culture of the strain, named QTYC-44, and storing it in the preservation medium (the formula is: Malt 20g, sucrose 2...
Embodiment 2
[0026] Embodiment 2, the identification of fungus QTYC-44
[0027] 1. Morphological identification
[0028] The preliminary morphological identification of the strain was based on the "Handbook of Ignorance of Ignorance Fungi". Such as figure 1 As shown, the colony grows faster on the MA (malt juice agar) medium, and the diameter of the colony is 60-65mm in 3 days under 28°C dark condition, and white conidia clusters can be seen near the edge of the colony. The clusters of conidia are densely arranged, and do not produce diffuse pigment and have no obvious odor. Conidiophores directly produce phialides or produce secondary branches, which tend to be opposite. Conidia are oval, light green, with smooth walls. The strain can grow in a test tube with Xinhua filter paper as the only carbon source, and the filter paper disintegrates obviously within 5 days, indicating that it has high biomass degradation activity.
[0029] 2. Molecular biological identification of strains
[0...
Embodiment 3
[0035] Embodiment 3, the mensuration of cellulase activity (CMC enzyme activity)
[0036] Inoculate 6 pieces of Trichodermasp.QTYC-44 bacteria cake (6mm in diameter) into 250ml of fermentation medium (recipe: 20g of malt, 20g of sucrose, 1g of peptone, 1L of distilled water, pH 7.0, sterilized at 120°C for 20min), Under the conditions of 28° C. and pH 7.0, ferment for 6 days, and take a small amount of fermentation liquid to measure the cellulase activity of the fermentation liquid. The fermentation broth was centrifuged at 4500r / min for 20min, and the supernatant was used as crude enzyme solution. Take 4 test tubes, 1 tube is used as blank control, 3 tubes are used as parallel sample tubes, add 1mL crude enzyme solution to each sample tube, preheat in 50℃ water bath for 2min, and then add 4mL crude enzyme solution to 3 test tubes respectively. Substrate solution preheated to 50°C (the preparation method of substrate solution is: accurately weigh 0.625g carboxymethylcellulose...
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