Repositorio UFRO · Omeka S

Physical Pretreatments Applied in Three Commercial Kits for the Extraction of High-Quality DNA from Activated Sewage Sludge

Primer Autor	Cid-Alda, Fernanda P.
Co-autores	Vasquez, Claudio Leyton-Carcaman, Benjamin Segovia, Inaky Pinto, Fernanda Abanto, Michel
Título	Physical Pretreatments Applied in Three Commercial Kits for the Extraction of High-Quality DNA from Activated Sewage Sludge
Editorial	MDPI
Revista	INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
Lenguaje	en
Resumen	Obtaining sufficient and high-quality genomic DNA from sludge samples is a fundamental issue of feasibility and comparability in genomic studies of microbial diversity. Commercial kits for soil are often used for the extraction of gDNA from sludge samples due to the lack of specific kits. However, the evaluation of the performance of commercial kits for sludge DNA extraction is scarce and optimization of these methods to obtain a high quantity and quality of DNA is necessary, especially for downstream genomic sequencing. Sequential batch reactors (SBRs) loaded with lignocellulosic biomass are used for the synthesis of renewable resources such as levulinic acid (LA), adipic acid (AA), and polyhydroxyalkanoates (PHAs), and the biochemical synthesis of these compounds is conducted through the inoculation of microbes present in the residual activated sludge (AS) obtained from a municipal wastewater treatment plant. To characterize these microbes, the extraction of DNA from residual sewage sludge was conducted with three different commercial kits: Nucleospin (R) Soil from Macherey-Nagel, DNEasy (R) PowerSoil (R) from Qiagen, and E.Z.N.A.(R) Plant DNA Kit from Omega BIO-TEK. Nevertheless, to obtain the highest load and quality of DNA for next-generation sequencing (NGS) analysis, different pretreatments and different combinations of these pretreatments were used. The pretreatments considered were an ultrasonic bath and a temperature of 80 degrees C, together and separately with different incubation time periods of 30, 60, and 90 min. The results obtained suggest a significant improvement in the efficiency and quality of DNA extraction with the three commercial extraction kits when used together with the ultrasonic bath and 80 degrees C for 60 min. Here, we were able to prove that physical pretreatments are a viable alternative to chemical lysis for DNA extraction from complex samples such as sludge.
Fecha Publicación	2023
Tipo de Recurso	artículo original
doi	10.3390/ijms242015243
Formato Recurso	PDF
Palabras Claves	DNA extraction complex samples next-generation sequencing metagenomics microbiome bioreactor
Ubicación del archivo	http://dx.doi.org/10.3390/ijms242015243
Categoría OCDE	Bioquímica y biología molecular Química
Materias	Extracción de ADN muestras complejas secuenciación de próxima generación metagenómica microbioma biorreactor
Identificador del recurso (Mandatado-único)	artículo original
Versión del recurso (Recomendado-único)	versión publicada
License	CC BY 4.0
Condición de la licencia (Recomendado-repetible)	CC BY 4.0
Derechos de acceso	acceso abierto
Access Rights	acceso abierto
Id de Web of Science	WOS:001095441200001
ISSN	1661-6596
Tipo de ruta	verde# dorado
Categoría WOS	Bioquímica y biología molecular Química
Referencia del Financiador (Mandatado si es aplicable-repetible)	ANID-FONDECYT 3210626 ANID-FONDEF ID19I10299 ANID 21210454 ANID 3200728 ANID FONDECYT 3210626 ANID FONDEF ID19I10299 UFRO PIA21-0002