Data/hora: 16/06/2024 06:25:29

Provedor de dados: | 5
País: | France
Título: | PCR performance of a thermostable heterodimeric archaeal DNA polymerase
Autores: | Killelea, Tom; Ralec, Celine; Bosse, Audrey; Henneke, Ghislaine.
Data: | 2014-05
Ano: | 2014
Palavras-chave: | DNA polymerase; Archaea; Family D; PCR; Pyrococcus.
Resumo: | DNA polymerases are versatile tools used in numerous important molecular biological core technologies like the ubiquitous polymerase chain reaction (PCR), cDNA cloning, genome sequencing, and nucleic acid based diagnostics. Taking into account the multiple DNA amplification techniques in use, different DNA polymerases must be optimized for each type of application. One of the current tendencies is to reengineer or to discover new DNA polymerases with increased performance and broadened substrate spectra. At present, there is a great demand for such enzymes in applications, e.g., forensics or paleogenomics. Current major limitations hinge on the inability of conventional PCR enzymes, such as Taq, to amplify degraded or low amounts of template DNA. Besides, a wide range of PCR inhibitors can also impede reactions of nucleic acid amplification. Here we looked at the PCR performances of the proof-reading D-type DNA polymerase from P abyssi, Pab-polD. Fragments, 3 kilobases in length, were specifically PCR-amplified in its optimized reaction buffer. Pab-polD showed not only a greater resistance to high denaturation temperatures than Taq during cycling, but also a superior tolerance to the presence of potential inhibitors. Proficient proof-reading Pab-polD enzyme could also extend a primer containing up to two mismatches at the 3' primer termini. Overall, we found valuable biochemical properties in Pab-polD compared to the conventional Taq, which makes the enzyme ideally suited for cutting-edge PCR-applications.
Tipo: | Text
Idioma: | Inglês

Identificador:

http://archimer.ifremer.fr/doc/00193/30450/28871.pdf

DOI:10.3389/fmicb.2014.00195

Editor: | Frontiers Research Foundation
Relação: | http://archimer.ifremer.fr/doc/00193/30450/
Formato: | application/pdf
Fonte: | Frontiers In Microbiology (1664-302X) (Frontiers Research Foundation), 2014-05 , Vol. 5 , N. 195 , P. 1-11
Fonte: | 2014 Killelea, Ralec, Bosse and Henneke. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.