How can I optimize PCR conditions for AT-rich templates?
How can I optimize PCR conditions for AT-rich templates?
Some templates may have long AT-rich stretches that are hard to amplify under standard reaction conditions. The Plasmodium falciparum genome is about 80% AT, and regions flanking genes are often AT rich.
Polymerases recommended for GC-rich templates, such as EmeraldAmp GT PCR Master Mix, EmeraldAmp Max PCR master mixes, and PrimeSTAR GXL DNA Polymerase, are also suitable for AT-rich templates.
The advantage of having AT-rich templates is that a lower extension temperature can be used. For certain templates with AT content >80–85%, the extension temperature can be lowered from 72°C to 65–60°C. DNA replication at this reduced temperature appears to be reliable (Su et al. 1996).
References
Su, X. Z., et al. Reduced Extension Temperatures Required for PCR Amplification of Extremely A+T-rich DNA. Nucl Acids Res. 24, 1574–1575 (1996).