I have to isolate fungal and bacterial microbiomes from the soil and to test their pathogenicity on rice plants. I have to grow seeds and then at which seedling stages, I may perform pathogenicity test. ;I want a cheap and less time consuming protocol for pathogenicity test.
Once you have identified pure cultures conclusively. Take 100 seed sample and put them on an semi soft agar plate use a lower level of agar. Shake seed on to get the fungal material to adhere on semi soft agar to the seed with mycelium. Then plant the inoculated seeds with their identification in sand flats. Plant at 3 cm depth keep moist. Beside replication use randomization for proper experimental design. This can be done either in the greenhouse or a growth chamber for more control. Use an noninoculated lot as a control. Replicate treatments for statistical analysis 3 replicates of 100 seed sufficient for Seed Technologists. Take data on emergence and growth and describe symptoms that develop if any. If you are interested in more advanced symptoms make sure you have ability to maintain plastic tent which saturated humidity can be maintained for foliar symptoms. I would use 2-3 weeks in the sand to see seedling reaction. Then I would mist and keep moist for another week to see foliar root symptoms if any.
Dear Aqleem,
I would test the plant stage in which you are most likely to see infections in the field. If there is no information on that I would test multiple stages. You can try to use leaf cuttings to test in vitro, but this does not always represent what happens in a fully functioning plant. Furthermore, it could well be that your pathogens usually infect roots so you might also have to consider root infections.
Kind regards,
Silvia
Once you have identified pure cultures conclusively. Take 100 seed sample and put them on an semi soft agar plate use a lower level of agar. Shake seed on to get the fungal material to adhere on semi soft agar to the seed with mycelium. Then plant the inoculated seeds with their identification in sand flats. Plant at 3 cm depth keep moist. Beside replication use randomization for proper experimental design. This can be done either in the greenhouse or a growth chamber for more control. Use an noninoculated lot as a control. Replicate treatments for statistical analysis 3 replicates of 100 seed sufficient for Seed Technologists. Take data on emergence and growth and describe symptoms that develop if any. If you are interested in more advanced symptoms make sure you have ability to maintain plastic tent which saturated humidity can be maintained for foliar symptoms. I would use 2-3 weeks in the sand to see seedling reaction. Then I would mist and keep moist for another week to see foliar root symptoms if any.
Suresh K Malhotra You are so kind to appreciate my comments. I have followed your comments and our shared interests much to my delight. I am hoping you have a Happy Prosperous New Year. Sincerely, Paul Reed Hepperly
I have to work on rice seeds, It means I have to dip the rice seeds in fungal cultures, but the question is how many fungal spores or how much concentration of fungal spores is needed to full fill pathogenicity on rice seeds.The concentration of the fungal spores for each seeds would be different if i dip it in a fungal culture. Second questions is the randomization is very important in any experiment. How would I make the treamtents and control? What would be positive and negative control? All are questions which need to be consider before doing an experiment?
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