I just found the Platinum SuperFi II DNA Polymerase, which should simplify PCR protocol as it allows uniform annealing temperature of 60°C, but it should also have very high fidelity; >300× higher than Taq Pol, that should be even more than Q5, reported by NEB to have 280× higher fidelity than Taq Pol.
The SuperFi II DNA Polymerase should even allow amplification up to 40 kbp, while Q5 only up to 20 kbp.
This looks like we have new Queen in the HighFidelity DNA Pol area, don't we? Does somebody have experience with this enzyme?
The Double Helix is wrong: there is no mutual twisting of the two strands -- they run in parallel (of course they are antiparallel). I proposed a different model, it is described in the article
https://www.researchgate.net/publication/339106477_DNA_the_Double_Helix_or_the_Ribbon_Helix
Hey, Tomás!
I have never used the Q5 enzyme, but I have already used some high-fidelity enzymes from Thermo.
For me, SuperFi II is the best! The protocol is very simple and I amplified large fragments with high GC content without any problems.
Hi Flávia Caroline Gan ,
thanks for your answer. Do you use the annealing temperature of 60°C for all reactions? Is the amplification reliable? Any troubles?
Yes and no.
60°C is used for almost all reactions, but not all. For example, in my case, I amplified some genes for Gibson cloning, so my oligos have a high size (50/60bp). In these cases, the annealing step is not necessary and you just proceed to the extension step. The enzyme buffer permits annealing at 72°C. In all my reactions, I always had high amplification and I always followed the protocol parameters.
You just need to be careful with the correct design of your primers and with your DNA sample. Your sample needs to be as pure as possible, and you should use the amount recommended by the protocol in each reaction, which is 10ng if the plasmid gene is being amplified. When I used a bit more sample, the amplification didn't occur.
I do find it reliable, much more so than other enzymes I've used. We haven't had any problems with genes amplified with it and used in cloning.
- Badges
- Science topic
- Similar topics
- Philosophy
- Moral Philosophy
- Free Will