Abstract
Full-length mitochondrial cytochrome c oxidase I (COI) sequence information from lobster phyllosoma larvae can be difficult to obtain when DNA is degraded or fragmented. Primers that amplify smaller fragments are also more useful in metabarcoding studies. In this study, we developed and tested a method to design a taxon-specific mini-barcode primer set for marine lobsters. The shortest, most informative portion of the COI gene region was identified in silico, and a DNA barcode gap analysis was performed to assess its reliability as species diagnostic marker. Primers were designed, and cross-species amplification success was tested on DNA extracted from a taxonomic range of spiny-, clawed-, slipper- and blind lobsters. The mini-barcode primers successfully amplified both adult and phyllosoma COI fragments, and were able to successfully delimit all species analyzed. Previously published universal primer sets were also tested and sometimes failed to amplify COI from phyllosoma samples. The newly designed taxon-specific mini-barcode primers will increase the success rate of species identification in bulk environmental samples and add to the growing DNA metabarcoding toolkit.
Highlights
Holthuis (1991) provided a detailed systematic catalogue of most the marine lobsters known up to the early 1990s, based solely on the morphology of adult specimens
We developed and tested a method to design a taxon-specific mini-barcode primer set for marine lobsters
The top two mini-barcode fragments for each window length were selected for further analyses based on: (1) high mean Kimura 2-parameter (K2P) distance; (2) few zero pairwise non-conspecific distances; and (3) high proportion of clades shared between the neighbor-joining tree from the full-length DNA sequence alignment and the tree constructed using only data from selected windows
Summary
Holthuis (1991) provided a detailed systematic catalogue of most the marine lobsters known up to the early 1990s, based solely on the morphology of adult specimens. The traditional classification system used in the catalogue recognized the superfamilies; Nephropoidea (clawed lobsters), Palinuroidea (spiny and slipper lobsters), Eryonoidea (blind lobsters) and the living fossil Glypheoidea within the decapod suborder Macrura Reptantia [1]. Chan (2010) updated the list of valid species by adding several newly described taxa, and organizing all living marine lobsters into four infraorders: Astacidea, Glypheidea, Achelata and Polychelida. The new (2010) checklist recognized six families, 55 genera and 248 species (with four subspecies) of marine lobsters [2]. Marine lobsters have cryptic early life-history stages. Phyllosomas are dorso-ventrally flattened, leaf-like and transparent, and moult through a series of developmental stages of increasing size and morphological complexity [4].
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