School of Earth, Environmental, and Marine Sciences Faculty Publications and Presentations

An effective procedure for DNA isolation and voucher recovery from millimeter-scale copepods and new primers for the 18S rRNA and cytb genes

Document Type

Article

Publication Date

11-2014

Abstract

Many investigators need to determine whether individuals belong to the same species. DNA-sequence data have helped with this task, but current procedures of DNA isolation from millimeter-scale crustaceans, such as harpacticoid copepods, leave little to no voucher material for morphological analysis, and many procedures yield only enough DNA for a single amplification reaction. We therefore developed a DNA-isolation procedure that yielded essentially intact exoskeletons and sufficient DNA for multiple polymerase chain reactions. DNA-amplification success of our DNA-isolation procedure was relatively insensitive to (1) the length of preservation time from sample collection to DNA isolations and (2) the length of time the DNA was stored at − 20 °C after isolation. An additional benefit of our procedure is therefore that the DNA isolated is relatively stable. Primers available for the nuclear 18S rRNA gene and the mitochondrial cytochrome oxidase b (cytb) gene are known not to work for many harpacticoids. We therefore designed primers that would amplify and sequence an ~ 750-base-pair fragment of the 18S rRNA gene and others that would amplify and sequence an ~ 450-base-pair fragment of the cytb gene. Both primer sets worked for at least 12 harpacticoid families.

Comments

Copyright © 2014 Elsevier B.V. All rights reserved.

Publication Title

Journal of Experimental Marine Biology and Ecology

DOI

https://doi.org/10.1016/j.jembe.2014.06.016

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