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Cycle sequencing is a simple method in which successive rounds of denaturation, annealing, and extension in a thermal cycler result in linear amplification of extension products. The products are then injected into a capillary. All current Applied Biosystems DNA sequencing kits use cycle sequencing protocols. We have focused on two general categories of sequencing chemistry approaches: Dye Primer chemistry and Dye Terminator chemistry. Learn more about Sequencing Chemistries.

After the sequencing reaction, it is important to remove unincorporated dye terminators and salts that may compete for capillary electrophoretic injection. Unincorporated terminators can co-migrate with the sequencing template, resulting in basecalling errors, and excess salt translates to poor signal-to-noise ratios. Learn more about Sample Cleanup.

After performing the post-sequencing reaction purification, samples are ready for analysis on an Applied Biosystems capillary electrophoresis-based genetic analyzer. During capillary electrophoresis, the products of the cycle sequencing reaction are injected electrokinetically into capillaries filled with polymer. High voltage is applied so that the negatively charged DNA fragments move through the polymer in the capillaries toward the positive electrode. Learn more about Electrophoresis.

After electrophoresis, Data Collection software creates a Sample File of the raw data. Using downstream software applications, further data analysis is required to translate the collected color-data images into the corresponding nucleotide bases. Learn more about Data Analysis.