Geotrichum candidum and application thereof in greenhouse soil remediation
A greenhouse and soil remediation technology, which is applied in the field of agricultural microbes to achieve the effects of improving and activating soil, fast growth rate, and strong growth and reproduction ability
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Embodiment 1
[0029] Example 1: Isolation, screening and identification of Geotrichum Candidum XHS0030BCGMCCNO.9435
[0030] 1. Isolation and screening of strains
[0031] (1) Separation
[0032] The Geotrichum candidum used in the present invention is sampled and separated from typical greenhouse soils with serious salinization, sandy soil or protruding hardening in Korla, southern Xinjiang. A strain with a certain antagonistic ability to soil problems such as salinization and soil compaction. Use the traditional plate culture method to isolate the microorganisms in the soil layer, purify the strains by the plate streak method, and use different culture temperatures, pH values, and medium as enrichment conditions to screen out a batch of well-growing microbial strains, and select one of them The strain numbered XHS0030B.
[0033] Separation steps: according to the gradient dilution method, weigh 10g of soil samples in 90mL sterile saline, activate for 30min at 25°C and perform gradient ...
Embodiment 2
[0045] Example 2: Molecular level of Geotrichum Candidum XHS0030BCGMCCNO.9435
[0046] 1. PCR amplification of the ITSrDNA sequence of Geotrichum candidum and its sequencing
[0047] Pick a small amount of single colonies of the XHS0030B strain, put them into an EP tube filled with 25 μL of sterile water, boil at 100°C for 8-10 minutes, and then quickly put them into the ice-water mixture for 5 minutes. Centrifuge at 10000r / min for 5min, store at 4°C, and take the supernatant when used.
[0048] The construction of ITSrDNA gene sequence determination and phylogenetic tree thereof: extract the total DNA of bacterial strain according to routine method, carry out the PCR amplification of ITSrDNA section with deionized water with the dilution universal primer ITS1 and ITS4, primer design is as follows:
[0049] ITS1(F): 5'-TCCGTAGGTGAACCTGCGG-3'
[0050] ITS4(R): 5'-TCCTCCGCTTATTGATATGC-3'
[0051] The 50 μl reaction system contains: 5 μl of 10×PCR buffer, 20 pmol of each primer,...
Embodiment 3
[0054] Example 3: Preparation of greenhouse soil remediation agent
[0055] (1) Inoculation: prepare solid culture medium of four strains of Geotrichum candidum, Rhodopseudomonas palustris, Candida utilis and Mortierella alpina, perform strict aseptic operation after sterilization, and transfer from the inclined plane to the plate , cultured at 28°C for 3 days.
[0056] (2) Primary culture: Pick a single colony from the solid medium in step (1) and transfer it to a 50mL Erlenmeyer flask filled with liquid medium, perform aseptic operation, and culture at 25°C and 120r / min for 24h- 48h.
[0057] (3) Secondary fermentation: Inoculate the primary culture strains into a 500mL Erlenmeyer flask filled with liquid medium, perform aseptic operation, and culture at 25°C and 120r / min for 36h-72h.
[0058] (4) Compatibility of strain fermentation broth: 70-80 parts of Geotrichum candida secondary fermentation broth prepared in step (3), 30-50 parts of Rhodopseudomonas paluscans seconda...
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