PCR Amplification of up to 35-kb DNA with High Fidelity and High Yield from λ Bacteriophage Templates
Abstract
A target length limitation to PCR amplification of DNA has been identified and addressed. Concomitantly, the base-pair fidelity, the ability to use PCR products as primers, and the maximum yield of target fragment were increased. These improvements were achieved by the combination of a high level of an exonuclease-free, N-terminal deletion mutant of Taq DNA polymerase, Klentaq1, with a very low level of a thermostable DNA polymerase exhibiting a 3'-exonuclease activity (Pfu, Vent, or Deep Vent). At least 35 kb can be amplified to high yields from 1 ng of lambda DNA template.
- Publication:
-
Proceedings of the National Academy of Science
- Pub Date:
- March 1994
- DOI:
- 10.1073/pnas.91.6.2216
- Bibcode:
- 1994PNAS...91.2216B