Biocontrol pythium oligandrum and application
A technology of Pythium and biocontrol agents, applied in the application, biocide, microorganism and other directions, can solve limited problems, and achieve the effects of reducing environmental pollution, good application prospects, and simple cultivation conditions
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Embodiment 1
[0034] The separation of embodiment 1 biocontrol Pythium oligandrum
[0035] The specific process is as follows:
[0036] 1. Collection of soil samples
[0037] The soil at a depth of 10-20 cm was collected from the ginger disease field in Lijiazhai Village, Pingli Town, Laicheng District, Laiwu City, Shandong Province, and the soil was packed and marked and brought back to the laboratory for separation.
[0038]2. Isolation of Pythium oligandrum
[0039] (1) Take the collected soil samples and use a coarse sieve to remove stones and uncorrupted plant residues.
[0040] (2) Weigh 50 g of sieved soil, add 500 ml of water, stir for 5 minutes with a glass rod, immediately filter the supernatant through sieves with apertures of 150, 61, and 38 microns, and collect the filtrate.
[0041] (3) Rinse the 38-micron sieve with a small amount of water, and collect the residue filtered on the net in a conical flask.
[0042] (4) Take 5ml of the residue suspension and add it on the sel...
Embodiment 2
[0044] Molecular identification of embodiment 2 Pythium oligandrum GAQ1
[0045] After using the CTAB method to extract the genomic DNA of the five isolated strains, use Vazyme's MaxSuper-Fidelity DNA Polymerase high-fidelity enzyme and primer pair ITS1(5'-TCCGTAGGTGAACCTGCGG-3') / ITS4(5'-TCCTCCGCTTATTGATATGC-3') to amplify the ITS region of eukaryotes, sent to Shenggong Biological Company (Shanghai ) Co., Ltd. sequenced to obtain nucleotide sequence information, and submitted the obtained sequence to the GenBank database for BLAST analysis and comparison. It was found that a strain GAQ1 isolated may be Pythium oligandrum.
[0046] In order to further verify whether the bacterial strain is Pythium oligandrum, use Vazyme's The primer pair of MaxSuper-Fidelity DNA Polymeras high-fidelity enzyme and two housekeeping genes of Pythium TUBUF2(5'-CGGTAACAACTGGGCCAAGG-3') / TUBUR1(5'-CCTGGTACTGCTGGTACTC-3') and FM66(5'-TAGGATTTCAAGATCCTGC-3' ) / FM58(5'-CCACAAATTTCACTACATTGA-3') to amp...
Embodiment 3
[0057] Embodiment 3 Plate Antagonism Detection of Pythium oligandrum GAQ1
[0058] 1. Tablet confrontation test
[0059] (1) Pythium oligandrum GAQ1 and Pythium conglomerates grown on 10% V8 solid plates for 1 day were respectively punched along the edge of the colony with a puncher with a diameter of 6 mm. The preparation method of the 10% V8 solid medium is to centrifuge the V8 juice at 5000rpm for 10min, filter through four layers of gauze, take 1ml of the filtrate, add 99ml of water, 1.5g of agar, and sterilize the culture medium at 121°C for 20min .
[0060] (2) Inoculate a plate of Pythium conglomerates at the edge of a 10% V8 solid plate with a distance of 9 cm, and inoculate a plate of Pythium oligandrum GAQ1 at the other end of the plate, and culture it at 25° C.
[0061] (3) Cover the confrontation plate with 5ml 0.1% trypan blue before contacting the two strains, partially cover, and completely cover three different stages, rotate gently to ensure that the dye is ...
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