Stool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform

Abstract Many studies correlate changes in human gut microbiome with the onset of various diseases, mostly by 16S rRNA gene sequencing. Setting up the optimal sampling and DNA isolation procedures is crucial for robustness and reproducibility of the results. We performed a systematic comparison of s...

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Autores principales:	Petra Videnska, Kristyna Smerkova, Barbora Zwinsova, Vlad Popovici, Lenka Micenkova, Karel Sedlar, Eva Budinska
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2019
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/37d469f2166f4732bf37c9877781576e
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spelling	oai:doaj.org-article:37d469f2166f4732bf37c9877781576e2021-12-02T15:09:37ZStool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform10.1038/s41598-019-49520-32045-2322https://doaj.org/article/37d469f2166f4732bf37c9877781576e2019-09-01T00:00:00Zhttps://doi.org/10.1038/s41598-019-49520-3https://doaj.org/toc/2045-2322Abstract Many studies correlate changes in human gut microbiome with the onset of various diseases, mostly by 16S rRNA gene sequencing. Setting up the optimal sampling and DNA isolation procedures is crucial for robustness and reproducibility of the results. We performed a systematic comparison of several sampling and DNA isolation kits, quantified their effect on bacterial gDNA quality and the bacterial composition estimates at all taxonomic levels. Sixteen volunteers tested three sampling kits. All samples were consequently processed by two DNA isolation kits. We found that the choice of both stool sampling and DNA isolation kits have an effect on bacterial composition with respect to Gram-positivity, however the isolation kit had a stronger effect than the sampling kit. The proportion of bacteria affected by isolation and sampling kits was larger at higher taxa levels compared to lower taxa levels. The PowerLyzer PowerSoil DNA Isolation Kit outperformed the QIAamp DNA Stool Mini Kit mainly due to better lysis of Gram-positive bacteria while keeping the values of all the other assessed parameters within a reasonable range. The presented effects need to be taken into account when comparing results across multiple studies or computing ratios between Gram-positive and Gram-negative bacteria.Petra VidenskaKristyna SmerkovaBarbora ZwinsovaVlad PopoviciLenka MicenkovaKarel SedlarEva BudinskaNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 9, Iss 1, Pp 1-14 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Petra Videnska Kristyna Smerkova Barbora Zwinsova Vlad Popovici Lenka Micenkova Karel Sedlar Eva Budinska Stool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform
description	Abstract Many studies correlate changes in human gut microbiome with the onset of various diseases, mostly by 16S rRNA gene sequencing. Setting up the optimal sampling and DNA isolation procedures is crucial for robustness and reproducibility of the results. We performed a systematic comparison of several sampling and DNA isolation kits, quantified their effect on bacterial gDNA quality and the bacterial composition estimates at all taxonomic levels. Sixteen volunteers tested three sampling kits. All samples were consequently processed by two DNA isolation kits. We found that the choice of both stool sampling and DNA isolation kits have an effect on bacterial composition with respect to Gram-positivity, however the isolation kit had a stronger effect than the sampling kit. The proportion of bacteria affected by isolation and sampling kits was larger at higher taxa levels compared to lower taxa levels. The PowerLyzer PowerSoil DNA Isolation Kit outperformed the QIAamp DNA Stool Mini Kit mainly due to better lysis of Gram-positive bacteria while keeping the values of all the other assessed parameters within a reasonable range. The presented effects need to be taken into account when comparing results across multiple studies or computing ratios between Gram-positive and Gram-negative bacteria.
format	article
author	Petra Videnska Kristyna Smerkova Barbora Zwinsova Vlad Popovici Lenka Micenkova Karel Sedlar Eva Budinska
author_facet	Petra Videnska Kristyna Smerkova Barbora Zwinsova Vlad Popovici Lenka Micenkova Karel Sedlar Eva Budinska
author_sort	Petra Videnska
title	Stool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform
title_short	Stool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform
title_full	Stool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform
title_fullStr	Stool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform
title_full_unstemmed	Stool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform
title_sort	stool sampling and dna isolation kits affect dna quality and bacterial composition following 16s rrna gene sequencing using miseq illumina platform
publisher	Nature Portfolio
publishDate	2019
url	https://doaj.org/article/37d469f2166f4732bf37c9877781576e
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Stool sampling and DNA isolation kits affect DNA quality and bacterial composition following 16S rRNA gene sequencing using MiSeq Illumina platform

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