Bacteriophage for preventing and treating plant protection ralstonia solanacearum and pseudomonas solanacearum diseases and application thereof
A technology of Ralstonia solanacearum and Pseudomonas solanacearum, applied in the direction of virus/phage, bacteriophage, application, etc.
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Embodiment 1
[0016] Example 1 Screening and purification of phage 10RS306A
[0017] 1. Isolation of tomato Ralstonia solanacearum pathogen
[0018] Take tomato rhizomes suffering from tomato bacterial wilt disease in Xiamen, Fujian, and put them in a sterile mortar, wash the surface of the rhizomes with 50 mL of 75% ethanol, grind them in a sterile mortar, and then use an inoculation loop to dip into the grinding solution. Streak on the TTC plate of R. solanacearum identification medium, culture at 30°C for 24 hours, pick red colonies, and purify by streaking on the same selection medium, repeat the purification 3 times, and pick the strains with the same shape with an inoculation loop A single colony was taken and inserted into LB liquid medium, cultured at 30 °C for 12 h, mixed with 500 μL of bacterial liquid and 500 μL of 40% glycerol, and stored in a -80 °C refrigerator.
[0019] 2. Expanded culture of R. solanacearum
[0020] For the expanded culture of host bacteria, prepare 1 L of...
Embodiment 2
[0034] 1. Determination of the optimal multiplicity of infection (MOI) of phage 10RS306A against R. solanacearum
[0035] Take eight 100 mL fresh LB liquid medium and process them separately. Add 1 mL of 10 6 cfu / mL R. solanacearum bacteria liquid and 1 mL content is 10 9 pfu / mL phage liquid, add 1 mL content to 10 7 cfu / mL R. solanacearum bacteria liquid and 1 mL content is 10 9 pfu / mL phage liquid, add 1 mL content to 10 at the same time for treatment 3 8 cfu / mL R. solanacearum bacteria liquid and 1 mL content is 10 9 pfu / mL phage liquid, add 1 mL at the same time for treatment 4, the content is 10 8 cfu / mL R. solanacearum bacteria liquid and 1mL content is 10 8 pfu / mL phage liquid, add 1 mL content to 10 at the same time for treatment 5 8 cfu / mL R. solanacearum bacteria liquid and 1 mL content is 10 7 pfu / mL bacteriophage liquid, add 1 mL content to 10 8 cfu / mL R. solanacearum bacteria liquid and 1 mL content is 10 6 pfu / mL phage liquid, add 1mL content to 10 at t...
Embodiment 3
[0046] The cracking ability and control effect of embodiment 3 bacteriophage 10RS306A
[0047] 1. Determination of the lysing ability of phage 10RS306A on R. solanacearum
[0048] The phage 10RS306A was counted and diluted to 1×10 8 , 1×10 7 , 1×10 6 , 1×10 5 , 1×10 4 There are 5 concentrations of pfu / mL, and sterile water is used as the blank control, and three parallels are made for each concentration. Take 6 tubes of cultured Ralstia solanacearum liquid, calculate the content of Ralstia solanacearum by dilution plate coating method before treatment, add 1 mL of phage liquid of 5 concentrations and sterile water respectively, and then After culturing for 12 h at 150 r / min at ℃, the lysing ability of phage 10RS306A on R. solanacearum was determined by calculating the content of R. solanacearum after treatment by dilution plate spreading method. The results showed that phage 10RS306A had a better bactericidal effect on R. solanacearum, and the higher the concentration, t...
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