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https://hdl.handle.net/1822/81084
Título: | DNA extraction leads to bias in bacterial quantification by qPCR |
Autor(es): | Lima, Ângela Martins França, Ângela Maria Oliveira Sousa Muzny, Christina A. Taylor, Christopher M. Cerca, Nuno |
Palavras-chave: | Bacterial load quantification gDNA extraction efficiency qPCR gDNA yield Exogenous control Calibration curve qPCR reaction efficiency |
Data: | 14-Nov-2022 |
Editora: | Springer Nature |
Revista: | Applied Microbiology and Biotechnology |
Citação: | Lima, A., França, A., Muzny, C.A. et al. DNA extraction leads to bias in bacterial quantification by qPCR. Appl Microbiol Biotechnol 106, 7993–8006 (2022). https://doi.org/10.1007/s00253-022-12276-4 |
Resumo(s): | Quantitative PCR (qPCR) has become a widely used technique for bacterial quantification. The affordability, ease of experimental design, reproducibility, and robustness of qPCR experiments contribute to its success. The establishment of guidelines for minimum information for publication of qPCR experiments, now more than 10 years ago, aimed to mitigate the publication of contradictory data. Unfortunately, there are still a significant number of recent research articles that do not consider the main pitfalls of qPCR for quantification of biological samples, which undoubtedly leads to biased experimental conclusions. qPCR experiments have two main issues that need to be properly tackled: those related to the extraction and purification of genomic DNA and those related to the thermal amplification process. This mini-review provides an updated literature survey that critically analyzes the following key aspects of bacterial quantification by qPCR: (i) the normalization of qPCR results by using exogenous controls, (ii) the construction of adequate calibration curves, and (iii) the determination of qPCR reaction efficiency. It is primarily focused on original papers published last year, where qPCR was applied to quantify bacterial species in different types of biological samples, including multi-species biofilms, human fluids, and water and soil samples. |
Tipo: | Artigo |
URI: | https://hdl.handle.net/1822/81084 |
DOI: | 10.1007/s00253-022-12276-4 |
ISSN: | 0175-7598 |
Versão da editora: | https://link.springer.com/article/10.1007/s00253-022-12276-4 |
Arbitragem científica: | yes |
Acesso: | Acesso restrito UMinho |
Aparece nas coleções: | CEB - Publicações em Revistas/Séries Internacionais / Publications in International Journals/Series |
Ficheiros deste registo:
Ficheiro | Descrição | Tamanho | Formato | |
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document_55897_1.pdf Acesso restrito! | 1,52 MB | Adobe PDF | Ver/Abrir |