Optimised biomolecular extraction for metagenomic analysis of microbial biofilms from high-mountain streams

PeerJ
S. B. BusiT. J. Battin

Abstract

Glacier-fed streams (GFS) are harsh ecosystems dominated by microbial life organized in benthic biofilms, yet the biodiversity and ecosystem functions provided by these communities remain under-appreciated. To better understand the microbial processes and communities contributing to GFS ecosystems, it is necessary to leverage high throughput sequencing. Low biomass and high inorganic particle load in GFS sediment samples may affect nucleic acid extraction efficiency using extraction methods tailored to other extreme environments such as deep-sea sediments. Here, we benchmarked the utility and efficacy of four extraction protocols, including an up-scaled phenol-chloroform protocol. We found that established protocols for comparable sample types consistently failed to yield sufficient high-quality DNA, delineating the extreme character of GFS. The methods differed in the success of downstream applications such as library preparation and sequencing. An adapted phenol-chloroform-based extraction method resulted in higher yields and better recovered the expected taxonomic profile and abundance of reconstructed genomes when compared to commercially-available methods. Affordable and straight-forward, this method consistently recapitul...Continue Reading

References

Feb 11, 1993·Nucleic Acids Research·T Mukhopadhyay, J A Roth
Feb 1, 1996·Applied and Environmental Microbiology·J ZhouJ M Tiedje
May 1, 1997·Journal of Applied Microbiology·S J DancerD J Platt
Nov 5, 1999·Applied and Environmental Microbiology·D N MillerW C Ghiorse
May 7, 2003·Journal of Microbiological Methods·Michael HowelerLarry P Walker
Feb 7, 2004·Molecular Biotechnology·Peter RådströmCharlotta Löfström
Jun 24, 1950·Nature·S BANERJEE, B C HALDAR
Oct 26, 2007·Applied Microbiology and Biotechnology·Cora CarriggVincent O'Flaherty
Jun 10, 2011·Environmental Microbiology·Janet K JanssonJack A Gilbert
Sep 20, 2011·Applied and Environmental Microbiology·Kyoung-Ho Kim, Jin-Woo Bae
Jan 13, 2012·The ISME Journal·Katharina BesemerTom J Battin
Jan 1, 2006·CSH Protocols·Joseph Sambrook, David W Russell
May 17, 2012·Microbial Informatics and Experimentation·Torsten ThomasFolker Meyer
Jul 31, 2012·Plant Methods·Zhanguo Xin, Junping Chen
Mar 15, 2013·The ISME Journal·Linda WilhelmKatharina Besemer
Mar 29, 2013·Nature·Karen G LloydBo Barker Jørgensen
Sep 26, 2013·Methods in Enzymology·Hugo RoumePaul Wilmes
Oct 10, 2013·FEMS Microbiology Ecology·Tatiana A VishnivetskayaTullis C Onstott
Oct 15, 2013·Bioinformatics·Luis M Rodriguez-R, Konstantinos T Konstantinidis
Jan 17, 2014·Environmental Microbiology·Linda WilhelmTom J Battin
Oct 21, 2014·Environmental Microbiology·Meghan ChafeeSheri L Simmons
Dec 18, 2014·Genome Biology·Michael I LoveSimon Anders
Apr 17, 2015·BMC Microbiology·J Paul BrooksGregory A Buck
May 23, 2015·Science·Shinichi SunagawaPeer Bork
Oct 11, 2016·Nature Microbiology·Anna Heintz-BuschartPaul Wilmes
Jan 5, 2017·Cold Spring Harbor Protocols·Michael R Green, Joseph Sambrook
Sep 7, 2017·Proceedings of the National Academy of Sciences of the United States of America·Alexander M MilnerLee E Brown
Jan 18, 2018·Scientific Data·Benjamin J TullyJohn F Heidelberg
May 29, 2018·Nature Microbiology·Christian M K SieberJillian F Banfield

❮ Previous
Next ❯

Citations

Jun 3, 2021·GigaScience·Pedro QueirósPaul Wilmes
Sep 16, 2021·The ISME Journal·Stilianos FodelianakisTom J Battin

❮ Previous
Next ❯

Methods Mentioned

BETA
PCR
nucleic acid extraction
amplicon sequencing
whole
whole genome shotgun sequencing
enzymatic treatment
electrophoresis

Software Mentioned

CheckM
R
mOTU
MEM
MEGAHIT
DASTool
BWA
kmer
VizBin
MaxBin2

Related Concepts

Related Feeds

Biofilm & Infectious Disease

Biofilm formation is a key virulence factor for a wide range of microorganisms that cause chronic infections.Here is the latest research on biofilm and infectious diseases.