Proofreading DNA polymerase fusions offer several advantages for long-range PCR, including faster run times and higher fidelity compared with Taq-based enzymes. However, their use so far has been limited to amplification of small to mid-range targets. In this article, we present a modified protocol for using a DNA polymerase fusion to amplify genomic targets exceeding 20 kb in length. This procedure overcomes several limitations of Taq blends, which up until recently, were the only option for long-range PCR. With a proofreading DNA polymerase fusion, high-molecular-weight amplicon can be generated and analyzed in a single day, and a significant proportion is expected to be error-free.
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