Comparison of DNA extraction protocols for the analysis of gut microbiota in fishes

FEMS Microbiology Letters
Andrea M LarsenCovadonga R Arias

Abstract

This study investigated the impacts of bacterial DNA extraction methodology on downstream analysis of fish gut microbiota. Feces and intestine samples were taken from three sympatric freshwater fish species with varying diets. Samples were processed immediately (approximately 4 h after capture; fresh), stored at -20 °C for 15 days or preserved in RNAlater® reagent for 15 days. DNA was then extracted using two commercial kits: one designed for animal tissues and one specifically formulated for stool samples. Microbial community fingerprints were generated using ribosomal intergenic spacer analysis. Factors including diversity as depicted by band number, band intensity, repeatability and practicalities such as cost and time were considered. Despite significant differences in microbiota structure, results were similar between feces and intestine samples. Frozen samples were consistently outperformed by other storage methods and the stool kit typically outperformed the tissue kit. Overall, we recommend extraction of bacterial DNA from fresh samples using the stool kit for both sample types. If samples cannot be processed immediately, preservation in RNAlater® is preferred to freezing. Choice of DNA extraction method significantly i...Continue Reading

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Apr 24, 2013·FEMS Microbiology Ecology·Andrea LarsenCovadonga R Arias
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Feb 18, 2014·Journal of Applied Microbiology·A M LarsenC R Arias

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Citations

Oct 27, 2015·Frontiers in Immunology·Ana Montalban-ArquesJorge Galindo-Villegas
Jul 28, 2016·Journal of Microbiological Methods·Ali H D JanabiKenneth H McKeever
Sep 2, 2016·Frontiers in Microbiology·Eleonora SattinBarbara Cardazzo
Sep 29, 2018·Indian Journal of Microbiology·Chandni TalwarRam Krishan Negi
Aug 15, 2017·Journal of Microbiological Methods·Kimberly A LackeyMichelle K McGuire

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