Taq Polymerase Error Rate and Profile Determined by High Coverage Single Molecule Amplicon Sequencing

DNA polymerases have an innate error rate which is polymerase and DNA context specific. To evaluate the errors generated by Platinum Taq DNA Polymerase (Invitrogen) we used this polymerase in a PCR targeting part of the CASK gene. This was sequenced on a PacBio using many sequencing passes of the same molecule. When using an increasing minimum number of passes error rates eventually become asymptotic. At this point remaining errors are likely in the DNA molecule itself and hence generated by the polymerase during PCR. At minimum 10 passes to call a read-of-insert, sequencing coverage remains high (~9.6k times) and the polymerase specific error rate and mutational profile can be identified.