Currently I am working on single spore isolation method. some spores are germinating and some are non germinating.When i transfered the germinating spores into PDA they are contaminated with bacteria or yeast and other fungi. please suggest which antibiotic is best and its concentration . how can i reduce this contamination
You should add lactic acid 2.5% 2 drops when you plated PDA in petridish. Also please make sure your laminar air flow clean and sterile it (the laminar) with ultra violet first. do Anything with sterile condition
Thank you. Lactic acid is used for preserving fungal structures. i have to addd directly into prepared pda plate before transfering single spores
Dear,
Your question is somewhat unclear................ you are going to isolate the fungus by songle spore inoculation right? if so, then from where any environmental soil? if yes then you must be prepared serial dilution............... and made spread plate on a PDA or whatever other medium. Now you must pick spore developed from a separate single colony and transfer it in an new pda plate.............keeping properly sterile environment. i think no need to add any antibiotic of growth retardant on PDA medium.
Hi Devadatha,
As suggested by Monica, ensure you're working in a highly sterile environment (sterile laminar flow - clean with disinfectant and allow UV for about 15-30mins; sterilized apparatus - toothpicks, scapels, sterile water, etc). Lactic acid is nice to use but 2.5% may be too high and depending on the fungus, may affect growth - try 0.2% (2ml lactic acid per 1000ml of PDA). You may also try transferring the germinating spore onto a plate of water agar (15g or 20g agar powder per 1000ml of distilled water) and grow for just about 3-5 days before transferring unto PDA again. All these depends on the kind of fungus. Cheers!
thank u . i have to add laqctic acid before the preparation of Pda or after pouring into petripalate
Your problem can be resolved. Single spore isolation of fungi by using hand made glass needle from capillary tube. Also you have to do it in sterile conditions. See the link: http://www.fungaldiversity.org/fdp/sfdp/FD_2_47-63.pdf
Yeah i done it previously but encountered contamination with bacteria. so i started using Chloramphenicol 50microgram/ml eventhough iam encountering contamination with bacteria as well as Yeast and aspergillus n penicillium. So please suggest me how to overcome this problem
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