A Viability Quantitative PCR Dilemma: Are Longer Amplicons Better?
PMA
biofilm
microbiome
propidium monoazide
qPCR efficiency
v-qPCR
viability
vitality
Journal
Applied and environmental microbiology
ISSN: 1098-5336
Titre abrégé: Appl Environ Microbiol
Pays: United States
ID NLM: 7605801
Informations de publication
Date de publication:
12 02 2021
12 02 2021
Historique:
pubmed:
29
12
2020
medline:
22
12
2021
entrez:
28
12
2020
Statut:
ppublish
Résumé
The development of viability quantitative PCR (v-qPCR) has allowed for a more accurate assessment of the viability of a microbial sample by limiting the amplification of DNA from dead cells. Although valuable, v-qPCR is not infallible. One of the most limiting factors for accurate live/dead distinction is the length of the qPCR amplicon used. However, no consensus or guidelines exist for selecting and designing amplicon lengths for optimal results. In this study, a wide range of incrementally increasing amplicon lengths (68 to 906 base pairs [bp]) was used on live and killed cells of nine bacterial species treated with a viability dye (propidium monoazide [PMA]). Increasing amplicon lengths up to approximately 200 bp resulted in increasing quantification cycle (
Identifiants
pubmed: 33361365
pii: AEM.02653-20
doi: 10.1128/AEM.02653-20
pmc: PMC8090887
doi:
Substances chimiques
DNA, Bacterial
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
e0265320Subventions
Organisme : KU Leuven (Katholieke Universiteit Leuven)
ID : C24/17/086
Organisme : Fonds Wetenschappelijk Onderzoek (FWO)
ID : G0B2719N
Organisme : Fonds Wetenschappelijk Onderzoek (FWO)
ID : G091219N
Organisme : Fonds Wetenschappelijk Onderzoek (FWO)
ID : FWO-1SA5719N