Influence of salts and pcr inhibitors on the amplification capacity of three thermostable DNA polymerases

Katarina Davalieva, Georgi D. Efremov

Abstract


The full potential of PCR as a rapid DNA detection method, is limited by inhibition of thermostable DNA polymerases by components in biological samples and substances used for purification of template DNA. We compared the inhibition effect of blood, phenol and various ions on Taq, Tth and Tne thermostable DNA polymerases prepared by us. The amplification capacity of DNA polymerases was tested on the amplification of a 631 bp fragment of a â- globin gene from a 100 ng DNA template under the optimal PCR buffer. Blood above 1 % (v/v) and phenol above 0.1 % (v/v) inhibited Taq, while Tne and Tth tolerated 10 to 30 times more. The inhibitory effect of ions is lowest for the Tth DNA polymerase, followed by Taq and Tne DNA polymerase. We conclude that the PCR inhibiting effect of some substances to Taq DNA polymerase can be eliminated by the use of a more resistant thermostable DNA polymerase, such as Tth or Tne DNA polymerases.


Keywords


Thermostable DNA polymerase; PCR; PCR-inhibitors

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DOI: http://dx.doi.org/10.20450/mjcce.2010.173

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