Synthesis and error correction methods in gene fabrication

Abstract

Gene Fabrication technology involves the development and optimization of methods relevant to the in vitro synthesis of any given target gene sequence(s) in the absence of template. The driving purpose of this field of research is to bring about the capability for on-demand fabrication of a DNA construct of arbitrary length and sequence quickly, efficiently, and cost-effectively. The first part of this document describes many of the important considerations in performing successful de novo gene synthesis from a survey of the literature as well as from our own work. Recommendations are made for a universally effective, robust, and simple protocol for potential users of gene synthesis, discussing important factors such as choice of protocol, source of commercial oligonucleotides, and polymerase choice. The second part of this document focuses on error correction. Reducing error rates is one of the main challenges in gene fabrication because high error rates preclude the possibility of fabricating long gene targets in a practical and economical manner. Improvements in error rates are essential for continued progress in the development of gene fabrication technology. I discuss the importance of error rate in gene synthesis from a practical standpoint and show results in the development of novel methods for the removal of errors from a pool of synthesized DNA.