NDLI: Comparison of 6 DNA extraction methods for isolation of high yield of high molecular weight DNA suitable for shotgun metagenomics Nanopore sequencing to detect bacteria

Content Provider	Springer Nature : BioMed Central
Author	Gand, Mathieu Bloemen, Bram Vanneste, Kevin Roosens, Nancy H. C. De Keersmaecker, Sigrid C. J.
Abstract	Background Oxford Nanopore Technologies (ONT) offers an accessible platform for long-read sequencing, which improves the reconstruction of genomes and helps to resolve complex genomic contexts, especially in the case of metagenome analysis. To take the best advantage of long-read sequencing, DNA extraction methods must be able to isolate pure high molecular weight (HMW) DNA from complex metagenomics samples, without introducing any bias. New methods released on the market, and protocols developed at the research level, were specifically designed for this application and need to be assessed. Results In this study, with different bacterial cocktail mixes, analyzed as pure or spiked in a synthetic fecal matrix, we evaluated the performances of 6 DNA extraction methods using various cells lysis and purification techniques, from quick and easy, to more time-consuming and gentle protocols, including a portable method for on-site application. In addition to the comparison of the quality, quantity and purity of the extracted DNA, the performance obtained when doing Nanopore sequencing on a MinION flow cell was also tested. From the obtained results, the Quick-DNA HMW MagBead Kit (Zymo Research) was selected as producing the best yield of pure HMW DNA. Furthermore, this kit allowed an accurate detection, by Nanopore sequencing, of almost all the bacterial species present in a complex mock community. Conclusion Amongst the 6 tested methods, the Quick-DNA HMW MagBead Kit (Zymo Research) was considered as the most suitable for Nanopore sequencing and would be recommended for bacterial metagenomics studies using this technology.
Related Links	https://bmcgenomics.biomedcentral.com/counter/pdf/10.1186/s12864-023-09537-5.pdf
Ending Page	14
Page Count	14
Starting Page	1
File Format	HTM / HTML
ISSN	14712164
DOI	10.1186/s12864-023-09537-5
Journal	BMC Genomics
Issue Number	1
Volume Number	24
Language	English
Publisher	BioMed Central
Publisher Date	2023-08-04
Access Restriction	Open
Subject Keyword	Life Sciences Microarrays Proteomics Animal Genetics and Genomics Microbial Genetics and Genomics Plant Genetics and Genomics DNA extraction Method evaluation HMW DNA Nanopore Metagenomics Long-read
Content Type	Text
Resource Type	Article
Subject	Biotechnology Genetics
Journal Impact Factor	3.5/2023
5-Year Journal Impact Factor	4.1/2023

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