Proteus mirabilis phage and application thereof
A technology of Proteus mirabilis and bacteriophage, applied in the direction of phage, virus/phage, application, etc., can solve the problems of poor heat resistance, achieve the effects of preventing and controlling food pollution, high safety, and wide range of acid-base tolerance
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Embodiment 1
[0037] Sewage samples were collected from cattle farms in many cities and counties in Guangxi Zhuang Autonomous Region, and fecal swabs were collected from pig farms. After the sewage was left to settle naturally, the upper layer liquid was taken. After centrifugation at 5000r / min at 4°C for 10 minutes, the supernatant was taken, and repeated three times. Filter with a 0.45 μm sterile filter, and then filter with a 0.22 μm sterile filter, and store the sewage filtrate in a refrigerator at 4°C for later use; use SS solid medium to isolate Proteus mirabilis Pm05 from the stool sample, which is the host bacterial liquid, and freeze it in -80℃, standby.
[0038]Inoculate the isolated Proteus mirabilis from pigs into LB liquid medium, culture in a constant temperature shaker at 37°C for 6 hours, save the cultured bacterial solution for backup, and mix it with the above-mentioned sewage filtrate stored in a refrigerator at 4°C. Add an equal volume of 2×LB liquid medium, shake and cu...
Embodiment 2
[0043] Enrichment of Proteus mirabilis phage vB_PmiP_pPm05
[0044] Add 2 mL of the fresh host bacteria solution in Example 1 and the spare phage stock solution in Example 1 to 5 mL of LB liquid medium, put it into a constant temperature shaker at 37°C and 180r / min to cultivate only the culture medium is clear, 6h, Centrifuge at 4°C and 8000rpm for 5min, take the supernatant, filter with a 0.22μm sterile filter, and the filtrate is the phage lysate. Add DNaseI and RNase A to the phage lysate to a final concentration of 1 μg / ml, incubate at 37°C for 30 minutes, add NaCl with a final concentration of 1M and bathe in ice for 1 hour (that is, add sodium chloride to make the final concentration of sodium chloride in the mixed solution The concentration is 1M), centrifuged at 4°C and 8000rpm for 10min, took the supernatant and added PEG 8000 with a volume concentration of 10%, overnight at 4°C, then centrifuged at 4°C and 8000rpm for 10min, discarded the supernatant, inverted for 5m...
Embodiment 3
[0047] Transmission Electron Microscopy Observation of Proteus mirabilis Phage
[0048] The concentrated phage suspension obtained in Example 2 was observed under an electron microscope, and 10 μL of the concentrated phage liquid was absorbed on the copper grid for 10 minutes, and the phage liquid was blotted from the periphery of the copper grid with filter paper, and the surface of the carbon film of the copper grid was covered with 2% phosphotungstic acid. The solution (pH 6.8) was dyed on the droplet for 10 min, and the dye solution was blotted from the periphery of the copper grid with filter paper, and dried naturally; then the prepared sample copper grid was observed with a transmission electron microscope, and the results were as follows: figure 2 Shown: the head of the phage is flat oval, the length of the head is about 145nm, and the length of the tail is about 10nm. According to the regulations of the International Committee on Taxonomy of Viruses, the phage belongs...
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