In terms of results and efficiency, can we find differences between classical PCR such as
Enzyme Activation: 95°C 10 min
PCR (40 cycles): 95°C 15 sec + 60°C 1 min
and a fast PCR?
Enzyme Activation: 95°C 20 sec
PCR (40 cycles): 95°C 3 sec 60°C 30 sec
Thanks for your help
Hi Virginie
Classical PCR and fast PCR are basically similar in terms of results and efficiency. ....the difference is the speed of the reaction.......one should use optimized enzymes and PCR mix for the fast PCR reaction. The advantage you get is quick results...
Hi Virginie
Classical PCR and fast PCR are basically similar in terms of results and efficiency. ....the difference is the speed of the reaction.......one should use optimized enzymes and PCR mix for the fast PCR reaction. The advantage you get is quick results...
It's true that you should optimize your own PCR mix and program. It's also a good idea to look at the enzyme producer's data concerning e.g. its recommended activation time. I use a BioLine Sybr mix with enzyme activation time of ~2 min for normal PCR.
In some thermocyclers the ramping speed differs for 'slow' and 'fast' PCR programs - I always use the fast ramping speed, as it saves a bit of time and still works perfectly fine for me.
If you're looking for quick DNA/RNA amplification, have you thought of trying Recombinase Polymerase Amplification (RPA)? RPA is analogous to PCR (ie uses two opposing primers, they're just a bit longer), just everything is done with enzymes, so it all runs at ~37C, rather than thermal cycling. Reactions tend to take 10-15 minutes and sensitivity, yields etc are similar to PCR. The down side is it's not long enough to have a decent coffee break whilst your reaction runs. Check out http://www.twistdx.co.uk/our_technology/
it depends to your PCR fragment size I have don ultra rapid PCR with conventional thermo cylers under 30 minuets with taq polymrase the reaction was 95 for 10 sec, 54 for 5 sec and 72 for 10 sec in 2o cycles
you can read my article in :
Development of an ultra rapid and simple multiplex polymerase chain reaction technique for detection of Salmonella typhi.
by Karami Ali, Ahmadi Zeynab, Safiri Zahra, Khalilpour Akbar, Morrovati Saeid
Saudi Medical Journal (2006)
Volume: 27, Issue: 8, Pages: 1134-1138
Hot off the press, here's a paper from a group in Göttingen, Germany, using RPA to detect Francisella tularensis in
Ordinary PCR test consist extraction of DNA ( or not using direct PCR on sample that's very difficult indeed specially some very complex samples like Enviro or clinical) then setting PCR reaction, PCR ,and post PCR like electrophoresis So when I talk about fast PCR I mean Shortest possible time from getting sample to the result in Normal PCR takes around 1 day but in my short PCR or fast or ultra fast it should take only 1 hr so You should speed up or optimize all stages from DNA extraction to Do electrophoresis I have done all of these and in some of my test I can do PCR from sample to result on 45 minutes
This trail may be outdated. If I want to amplify genes of 3 to 16kb, what is the fastest way to do that?
There is no difference in the efficiency and result (product), but just reduce time of running also should be adjust the mixing of enzyme and ingredient
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