I have purchased "sigma direct PCR kit" and have used it. it was given to me with its own buffer and polymerase. Can anyone tell me, whether the kit can be used with normal PCR buffer and Taq? Does the buffer given with the kit has specific composition, and what is the composition?
PCR buffer composition: grade water for PCR , Tris-HCl, and KCl , adding MgCl2 according to PCR protocol. This solution is suitable condition for activity of Taq enzyme.
The PCR buffer is chosen based on the type of polymerase is used in the reaction. It is possible to use Taq from a different company along with its suitable buffer. However, the quality of template has to be kept in mind. Does the direct kit allows amplification of DNA directly from tissues or cells. In that case the kit may contain DNA release reagents which may influence the reaction. The DNA polymerases provided in such kits are modified to improve their fidelity. You can still use other polymerases however, it is possible that the PCR efficiency of this reaction may be affected. If you choose to use other polymerases and buffers you may have to optimize the reaction. Make sure you use a hot start DNA polymerase. Optimize annealing temperature for your primer pair using the Tm Calculator provided by Sigma to when using Taq or any other polymerase. You can also do a gradient PCR to estimate most optimal anealing temperature with the DNA polymerase of your choice.
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