Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species
Bacteria containing mycolic acids in their cell envelope are often recalcitrant to cell lysis, so extracting DNA of sufficient quality for third-generation sequencing and high-fidelity genome assembly requires optimization, even when using commercial kits with protocols for hard-to-lyse bacteria. We...
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ftunivtasecite:oai:ecite.utas.edu.au:145365 2023-05-15T13:42:41+02:00 Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species Nahar, A Baker, AL Nichols, DS Bowman, JP Britz, ML 2021 application/pdf https://doi.org/10.3390/microorganisms9061253 http://www.ncbi.nlm.nih.gov/pubmed/34207615 http://ecite.utas.edu.au/145365 en eng MDPI http://ecite.utas.edu.au/145365/1/145365 - Benchmarking DNA extraction methods for phylogenomic analysis of sub-antarctic Rhodococcus and Williamsia species.pdf http://dx.doi.org/10.3390/microorganisms9061253 Nahar, A and Baker, AL and Nichols, DS and Bowman, JP and Britz, ML, Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species, Microorganisms, 9, (6) Article 1253. ISSN 2076-2607 (2021) [Refereed Article] http://www.ncbi.nlm.nih.gov/pubmed/34207615 http://ecite.utas.edu.au/145365 Biological Sciences Microbiology Microbial genetics Refereed Article PeerReviewed 2021 ftunivtasecite https://doi.org/10.3390/microorganisms9061253 2022-12-12T23:17:07Z Bacteria containing mycolic acids in their cell envelope are often recalcitrant to cell lysis, so extracting DNA of sufficient quality for third-generation sequencing and high-fidelity genome assembly requires optimization, even when using commercial kits with protocols for hard-to-lyse bacteria. We benchmarked three spin-column-based kits against a classical DNA extraction method employing lysozyme, proteinase K and SDS for six lysozyme-resistant, sub-Antarctic strains of Corynebaceriales . Prior cultivation in broths containing glycine at highly growth-inhibitory concentrations (4.04.5%) improved cell lysis using both classical and kit methods. The classical method produced DNA with average fragment sizes of 2759 Kbp and tight fragment size ranges, meeting quality standards for genome sequencing, assembly and phylogenomic analyses. By 16S rRNA gene sequencing, we classified two strains as Williamsia and four strains as Rhodococcus species. Pairwise comparison of average nucleotide identity (ANI) and alignment fraction (AF), plus genome clustering analysis, confirmed Rhodococcus sp. 1163 and 1168 and Williamsia sp. 1135 and 1138 as novel species. Phylogenetic, lipidomic and biochemical analyses classified psychrotrophic strains 1139 and 1159 as R. qingshengii and R. erythropolis , respectively, using ANI similarity of >98% and AF >60% for species delineation. On this basis, some members of the R. erythropolis genome cluster groups, including strains currently named as R. enclensis , R. baikonurensis , R. opacus and R. rhodochrous , would be reclassified either as R. erythropolis or R. qingshengii . Article in Journal/Newspaper Antarc* Antarctic eCite UTAS (University of Tasmania) Antarctic Microorganisms 9 6 1253 |
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Biological Sciences Microbiology Microbial genetics |
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Biological Sciences Microbiology Microbial genetics Nahar, A Baker, AL Nichols, DS Bowman, JP Britz, ML Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species |
topic_facet |
Biological Sciences Microbiology Microbial genetics |
description |
Bacteria containing mycolic acids in their cell envelope are often recalcitrant to cell lysis, so extracting DNA of sufficient quality for third-generation sequencing and high-fidelity genome assembly requires optimization, even when using commercial kits with protocols for hard-to-lyse bacteria. We benchmarked three spin-column-based kits against a classical DNA extraction method employing lysozyme, proteinase K and SDS for six lysozyme-resistant, sub-Antarctic strains of Corynebaceriales . Prior cultivation in broths containing glycine at highly growth-inhibitory concentrations (4.04.5%) improved cell lysis using both classical and kit methods. The classical method produced DNA with average fragment sizes of 2759 Kbp and tight fragment size ranges, meeting quality standards for genome sequencing, assembly and phylogenomic analyses. By 16S rRNA gene sequencing, we classified two strains as Williamsia and four strains as Rhodococcus species. Pairwise comparison of average nucleotide identity (ANI) and alignment fraction (AF), plus genome clustering analysis, confirmed Rhodococcus sp. 1163 and 1168 and Williamsia sp. 1135 and 1138 as novel species. Phylogenetic, lipidomic and biochemical analyses classified psychrotrophic strains 1139 and 1159 as R. qingshengii and R. erythropolis , respectively, using ANI similarity of >98% and AF >60% for species delineation. On this basis, some members of the R. erythropolis genome cluster groups, including strains currently named as R. enclensis , R. baikonurensis , R. opacus and R. rhodochrous , would be reclassified either as R. erythropolis or R. qingshengii . |
format |
Article in Journal/Newspaper |
author |
Nahar, A Baker, AL Nichols, DS Bowman, JP Britz, ML |
author_facet |
Nahar, A Baker, AL Nichols, DS Bowman, JP Britz, ML |
author_sort |
Nahar, A |
title |
Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species |
title_short |
Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species |
title_full |
Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species |
title_fullStr |
Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species |
title_full_unstemmed |
Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species |
title_sort |
benchmarking dna extraction methods for phylogenomic analysis of sub-antarctic rhodococcus and williamsia species |
publisher |
MDPI |
publishDate |
2021 |
url |
https://doi.org/10.3390/microorganisms9061253 http://www.ncbi.nlm.nih.gov/pubmed/34207615 http://ecite.utas.edu.au/145365 |
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Antarctic |
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Antarctic |
genre |
Antarc* Antarctic |
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Antarc* Antarctic |
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http://ecite.utas.edu.au/145365/1/145365 - Benchmarking DNA extraction methods for phylogenomic analysis of sub-antarctic Rhodococcus and Williamsia species.pdf http://dx.doi.org/10.3390/microorganisms9061253 Nahar, A and Baker, AL and Nichols, DS and Bowman, JP and Britz, ML, Benchmarking DNA extraction methods for phylogenomic analysis of sub-Antarctic Rhodococcus and Williamsia species, Microorganisms, 9, (6) Article 1253. ISSN 2076-2607 (2021) [Refereed Article] http://www.ncbi.nlm.nih.gov/pubmed/34207615 http://ecite.utas.edu.au/145365 |
op_doi |
https://doi.org/10.3390/microorganisms9061253 |
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Microorganisms |
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