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Open AccessArticle

Pretreatment Method for DNA Barcoding to Analyze Gut Contents of Rotifers

1
Department of Environmental Science and Engineering, Kyung Hee University, Yongin 17104, Korea
2
Department of Bioscience, Aarhus University, Vejlsøvej 25, 8600 Silkeborg, Denmark
3
Nakdong River Environment Research Center, National Institute of Environmental Research, Goryeong 40103
4
Department of Integrated Biological Science, Pusan National University, Busan 46241, Korea
5
Faculty of Marine Technology, Chonnam National University, Yeosu 59626, Korea
6
Fisheries Science Institute, Chonnam National University, Yeosu 59626, Korea
*
Authors to whom correspondence should be addressed.
Appl. Sci. 2020, 10(3), 1064; https://doi.org/10.3390/app10031064
Received: 29 December 2019 / Revised: 25 January 2020 / Accepted: 26 January 2020 / Published: 5 February 2020
(This article belongs to the Special Issue Food Chains and Food Webs in Aquatic Ecosystems)
We designed an experiment to analyze the gut content of Rotifera based on DNA barcoding and tested it on Asplanchna sp. in order to ensure that the DNA extracted from the rotifer species is from the food sources within the gut. We selected ethanol fixation (60%) to minimize the inflow effects of treated chemicals, and commercial bleach (the final concentration of 2.5%, for 210 s) to eliminate the extracellular DNA without damage to the lorica. Rotifers have different lorica structures and thicknesses. Therefore, we chose a pretreatment method based on Asplanchna sp., which is known to have weak durability. When we used the determined method on a reservoir water sample, we confirmed that the DNA fragments of Chlorophyceae, Diatomea, Cyanobacteria, and Ciliophora were removed. Given this result, Diatomea and cyanobacteria, detected from Asplanchna, can be considered as gut contents. However, bacteria were not removed by bleach, thus there was still insufficient information. Since the results of applying commercial bleach to rotifer species confirmed that pretreatment worked effectively for some species of rotifers food sources, in further studies, it is believed to be applicable to the gut contents analysis of more diverse rotifers species and better DNA analysis techniques by supplementing more rigorous limitations. View Full-Text
Keywords: gut content of Rotifera; eliminate the extracellular DNA; commercial bleach; pretreatment gut content of Rotifera; eliminate the extracellular DNA; commercial bleach; pretreatment
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Oh, H.-J.; Krogh, P.H.; Jeong, H.-G.; Joo, G.-J.; Kwak, I.-S.; Hwang, S.-J.; Gim, J.-S.; Chang, K.-H.; Jo, H. Pretreatment Method for DNA Barcoding to Analyze Gut Contents of Rotifers. Appl. Sci. 2020, 10, 1064.

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