I want a clear cut protocol regarding screening of biocontrol agents from the soil by using sclerotia of R. solani as baiting test. If someone has knowledge regarding this topic please do share your information with me.
To access the ability to identify biocontrol agents from soil. First identify a good medium and crop combination for your testing. Perhaps I would suggest potato and its Rhizoctonia would be a good system for study. I believe Rhizoctonia on potato dextrose agar would be suitable. The ability to get 400 to 1,000 scelerotia would provide 4 lots of 100 to 250 sclerotia each. A diverse sample of top soil from areas of long term potato production would have the antagonistic microflora. Your dry soil sample is used to treat your sclerotia I would leave the soil treated sclerotia for 1, 2, 4, and 8 days and then recover the sclerotia by surface disinfection by 10% bleach for 4 minutes and 1 min rinse in sterile distilled water. You then reisolate off the sclerotia and identify the invaders with a assay of viability and invaders. Once you have your sclerotia invaders identifed the isolation and identification will be followed by dual culture to gain more information on the interaction and its nature. Hope this is helpful.
I have dried soil for two weeks. I have also got scelrotia. Now What would be the next step Sir? In what depth, I bury the sclerotia and may I wrap sclerotia in a paper or just directly bury in the soil
Take the dry soil and put into glass petri plates. Moisten with sterile distilled water to moist but not supersaturated. Plant your scelerotia on the the soil plates. Harvest and surface disinfect the sclerotia after 1, 2, 4, and 8 days. Use 10% bleach as the surface disinfection on the retrieved scelerotia rise the 4 min disinfection with 1 min under sterile distilled water. Plate out your surface disinfected sclerotia onto a general media such as potato dextrose agar. Keep notes on the scelerotia germination and colonization of the scelerotia on viability of colonized scelerotia. Isolate purify and identify the fungi and bacteria that colonize the sclerotia from the soil. Grow pure culture scelerotia in dual culture with your colonist fungi and bacteria. Hope this is helpful.
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