Salmonella abortus equi phage and application thereof
A Salmonella and bacteriophage technology, applied in the field of bioengineering, can solve the problem of not finding bacteriophage, etc., and achieve the effects of reducing the occurrence and spread of equine abortion, strong lysis ability, and convenient use.
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Embodiment 1
[0023] Example 1 Screening and purification of Salmonella equine abortion phage
[0024] (1) Collection of samples
[0025] The donkey manure samples in the present invention were collected from a donkey farm in Dong'e County, Liaocheng City.
[0026] (2) Specific amplification of phages against Salmonella equine abortus in samples
[0027] Take 3 grams of sample and add it to 200 mL LB broth, then add 500 μL of Salmonella equine abortus, mix well and culture overnight at 37°C.
[0028] (3) Detection of the presence of phages by double-layer plate method
[0029] Prepare a nutrient agar solid medium with an agar content of 2%. After autoclaving, put it at room temperature to about 40-60°C, pour 10-15mL into a petri dish, spread it evenly on the bottom of the dish, and let it solidify at room temperature for 30 minutes. This nutrient agar was used as bottom agar. Take the above-mentioned mixed culture solution of Escherichia coli and the sample, centrifuge at 10000r / min for...
Embodiment 2
[0036] Example 2 Morphological observation of bacteriophage
[0037] Take 20 μL of phage proliferation solution and add it dropwise on the copper grid. After 15 minutes of precipitation, absorb the excess liquid with filter paper, add 15 μL of 2% phosphotungstic acid staining solution dropwise, stain for 5 minutes, absorb the excess dye solution, and observe it with an electron microscope after drying.
[0038] Electron microscope observation results are: phage DS2 is a long-tailed phage with a hexahedral symmetrical head with a diameter of about 75nm and a tail with a length of about 150nm. See figure 1 .
Embodiment 3
[0039] Example 3 Molecular biology identification of phage
[0040] 1. Gene sequencing of bacteriophage DS2
[0041] After the genome of bacteriophage DS2 was extracted, the whole genome was sequenced, and the sequencing result was: the full length of the genome was 114770bp, and the GC content was 40.26%. A total of 182 open reading frames and 21 tRNAs were compared by rast. Avirulence gene resistance gene.
[0042] Sequence similarity analysis using BLAST online tool (http: / / blast.ncbi.nlm.nih.gov / ), the results showed that the phage with the highest homology was Salmonella phage Sepoy, and its homology reached 97%. It shows that phage DS2 is a new strain of phage. From the sequencing results, it can be seen that the nucleic acid is a double-stranded linear DNA, which is a long-tailed bacteriophage.
[0043]In addition, according to the results of genome sequencing, it can be obtained that the sequence of the tail fiber protein gene sequence related to phage host recogni...
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