I need to figure the concentration of amplified DNA after a PCR by using available information like concentration of template DNA, enzyme speed, number of PCR cycles and so forth. Someone told me that i should be able to figure it out myself but im really bad with numbers and math so i got nowhere. Is there any software or some math equation or procedure that i can follow to the above mentioned effect?
Thanks a bunch
well a good rule of thumb is 2^n where n is number of cycles (provided you have a single dsDNA molecule to start with). so after 10 cycles e.g. 1024 molecules, but i don’t know if that is sufficient for you…
You could use a fluorometric dye such as Hoechst33258. It is rel. cheap (around 20$/100mg) and it specifically binds to dsDNA. It has a linear range from at least 50-800 ng DNA/mL. With special assays you can go down to 5 ng/mL.
Other options would be Ethidium bromide (which is around 10 times less sensitive) or the fluorescent dye SybrGreen.