PT - JOURNAL ARTICLE AU - Nathan B. Lubock AU - Di Zhang AU - George M. Church AU - Sriram Kosuri TI - A systematic comparison of error correction enzymes by next-generation sequencing AID - 10.1101/100685 DP - 2017 Jan 01 TA - bioRxiv PG - 100685 4099 - http://biorxiv.org/content/early/2017/01/15/100685.short 4100 - http://biorxiv.org/content/early/2017/01/15/100685.full AB - Gene synthesis, the process of assembling gene-length fragments from shorter groups of oligonucleotides (oligos), is becoming an increasingly important tool in molecular and synthetic biology. The length, quality, and cost of gene synthesis is limited by errors produced during oligo synthesis and subsequent assembly. Enzymatic error correction methods are cost-effective means to ameliorate errors in gene synthesis. Previous analyses of these methods relied on cloning and Sanger sequencing to evaluate their efficiencies, limiting quantitative assessment and throughput. Here we develop a method to quantify errors in synthetic DNA by next-generation sequencing. We analyzed errors in a model gene assembly and systematically compared six different error correction enzymes across 11 conditions. We find that ErrASE and T7 Endonuclease I are the most effective at decreasing average error rates (up to 5.8-fold relative to the input), whereas MutS is the best for increasing the number of perfect assemblies (up to 25.2-fold). We are able to quantify differential specificities such as ErrASE preferentially corrects C/G → G/C transversions whereas T7 Endonuclease I preferentially corrects A/T → T/A transversions. More generally, this experimental and computational pipeline is a fast, scalable, and extensible way to analyze errors in gene assemblies, to profile error correction methods, and to benchmark DNA synthesis methods.