PCR amplification of up to 35-kb DNA with high fidelity and high yield from λ bacteriophage templates

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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 λ DNA template.

Original languageEnglish
Pages (from-to)2216-2220
Number of pages5
JournalProceedings of the National Academy of Sciences of the United States of America
Volume91
Issue number6
DOIs
StatePublished - Mar 15 1994

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