Taq polymerase that can run 100 cycles? - (Sep/28/2005 )

Thanks, DNATI
But why did you select 40 cycles? Many parameters might affect the stabilities of Taq. For example, extension time, denature temperature (I use 92C instead of 94C), and so on.
Looking forward to your reply.

regards,

-Sanzhen

The major problem you face with doing large numbers of linear extension cycles is primer dimers. Even if your primers have only a low level of self homology then you will get primer dimer problems. Much better to just do multiple reactions and pool.

Daniel

DNA sequencing analysis software

Hi there,
Most Thermus aquaticus (Taq) polymerases have similar properties so you would want to change into something totally different like Archaeal hyperthermophiles like Pyrococcus furiosus (Pfu) or Thermococcus kodakaraensis (KOD) or Phusion (from NEB/Finnzymes) which have better stablilty than Taq. However the simplest thing to do and which I often do when faced with many cycles requirements, is that after 10 cycles, I lower the denaturation temp to 89°C or so and you usually get good results, since complex DNA does not reanneal very well and this temp is enough for denaturation of this kind

Good luck with your experiments
Dodobird