At-Rich Gene Synthesis

Over the last 15 years, the costs of oligonucleotide synthesis and gene synthesis have declined sharply and the practice of ordering these from commercial entities is now common lab protocol.  These companies also use synthetic oligonucleotides as building blocks for gene synthesis, making the synthesis of whole genes and larger constructs far more accessible.  However, there are limitations to which double-stranded DNA sequences they can synthesize with confidence.  These include ceilings on overall and local AT-content, homopolymers, palindromes, hairpins and repeats.  We report here improvements in the process of gene synthesis adapted for such difficult sequences. In this work, two genes from the AT-rich organism Plasmodium falciparum were synthesized from designed oligonucleotides using a polymerase construction and amplification method.  The component oligonucleotides were designed using DNAWorks and multiple polymerases were evaluated.