can anyone tell me my fungal sample is not getting amplified at all? I am using ITS as a universal primer and making master mix properly .this is my 5 th time can anyone tell the reason.
More information is needed Hitesh. DNA purification method and yield and its purity od260/280 would be helpful and amount used in the reaction. PCR conditions and primer annealing temperatures. Age of primers (approximate) and what they are stored in ( buffer,water and what temperature). Do you get a primer dimer or absolutely nothing or bands of the wrong size?. Do you have any other primers you can use as a control that at least something can amplify something from your dna
Details please.
thank you
paul
You need to use some controls. Do you have a DNA sample that you know has amplified with those specific primers? Primers are cheap so you can always order a fresh stock and/or design and buy more.
Positive control can help to detect the error. check your DNA integrity as well.
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