4.2.3 Cycle Sequencing

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 loaded onto a gel. See 4.2.5 for information on the used cycle sequencing protocols.

Advantages of Cycle Sequencing:

• Protocols are robust and easy to perform.
• Cycle sequencing requires much less template DNA than single-temperature extension methods.
• Cycle sequencing is more convenient than traditional single-temperature labelling methods that require a chemical denaturation step for double-stranded templates.
• High temperatures reduce secondary structure, allowing for more complete extension.
• High temperatures reduce secondary primer-to-template annealing.

© 2001 Alexander Binder