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Microbial Genomics logo Microbial Genomics

Volume 10, Issue 9

Evaluation of commercial RNA extraction kits for long-read metatranscriptomics in soil Open Access

Abstract

Metatranscriptomic analysis of the soil microbiome has the potential to reveal molecular mechanisms that drive soil processes regulated by the microbial community. Therefore, RNA samples must be of sufficient yield and quality to robustly quantify differential gene expression. While short-read sequencing technology is often favoured for metatranscriptomics, long-read sequencing has the potential to provide several benefits over short-read technologies. The ability to resolve complete transcripts on a portable sequencing platform for a relatively low capital expenditure makes Oxford Nanopore Technology an attractive prospect for addressing many of the challenges of soil metatranscriptomics. To fully enable long-read metatranscriptomic analysis of the functional molecular pathways expressed in these diverse habitats, RNA purification methods from soil must be optimised for long-read sequencing. Here we compare RNA samples purified using five commercially available extraction kits designed for use with soil. We found that the Qiagen RNeasy PowerSoil Total RNA Kit performed the best across RNA yield, quality and purity and was robust across different soil types. We found that sufficient sequencing depth can be achieved to characterise the active community for total RNA samples using Oxford Nanopore Technology, and discuss its current limitations for differential gene expression analysis in soil studies.

  • Received:
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Keyword(s): metatranscriptomics , Oxford Nanopore , RNA , soil , kit comparison and microbiome
Funding
This study was supported by the:
  • Shell United Kingdom (Award CW648947)
    • Principal Award Recipient: RichardK. Tennant
© 2024 The Authors
  • This is an open-access article distributed under the terms of the Creative Commons Attribution License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
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2024-09-19
2025-11-16

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Evaluation of commercial RNA extraction kits for long-read metatranscriptomics in soil
M Gen 10, 001298 (2024); https://doi.org/10.1099/mgen.0.001298
/content/journal/mgen/10.1099/mgen.0.001298
/content/journal/mgen/10.1099/mgen.0.001298
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