Cross-Site Evaluation of Commercial Sanger Sequencing Chemistries

Sanger sequencing remains an essential tool utilized by researchers. Despite competition from commercial sequencing providers, many academic sequencing core facilities continue to offer these services based on a model of competitive pricing, knowledgeable technical support, and rapid turnaround time. In-house Sanger sequencing remains a viable core service and, until recently, Applied Biosystems BigDye Terminator chemistry was the only commercially available solution for Sanger DNA sequencing on Applied Biosystems (ABI) instruments; however, several new products employing novel dye chemistries and reaction configurations have entered the market. As a result, there is a need to benchmark the performance of these new chemistries on various DNA templates, including difficult-to-sequence templates, and their amenability to commonly employed cost-saving measures, such as dye dilution and reaction miniaturization. To evaluate these new reagents, a study was designed to compare the quality of Sanger sequencing data produced by ABI BigDye and commercially available kits from 2 other vendors using both control and difficult-to-sequence DNA templates under various reaction conditions. This study will serve as a valuable resource to core facilities conducting Sanger sequencing that wish to evaluate the use of an alternative chemistry in their sequencing core.