Journal
3 BIOTECH
Volume 7, Issue -, Pages -Publisher
SPRINGER HEIDELBERG
DOI: 10.1007/s13205-017-0745-2
Keywords
De novo gene synthesis; High-fidelity polymerase; One-step PCR; Fel d 4
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Nowadays enzymatic synthesis of genes is the most powerful tool for fast resolution of the various tasks in the field of basic and applied biological research. PCR-based gene assembly from overlapping oligonucleotides has become a widely used strategy. However, all the methods described in the literature are not perfect and need an extra processing step. In this study we are verifying Phusion high-fidelity polymerase as a tool to reduce nucleotide mismatches in de novo gene synthesis, thus facilitating subsequent cloning. To test the efficiency of the polymerase, we selected Fel d 4 gene, which is a 581 bp DNA sequence encoding the lipocalin allergen protein, one of the major cat allergens. The approach described here, therefore, would be useful in DNA sequences creation.
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