NOTE Strange and Stepien: Perco flavescens and P. fluviotilis distinguished in fried fish fillets by DNA analysis 



293 



on a 2% agarose gel in TAE (Tris-acetate-EDTA, p.H. 

 8.0) buffer. The band was made visible with ethidium 

 bromide staining and was then excised and purified in 

 a spin column. The resulting gel-purified PCR product 

 was then used as a template for another round of PCR 

 with identical parameters. Samples consisting of 250 

 ng (nanograms) of purified PCR product and 16 pmol 

 (pico moles) of primer (L14724) were sequenced on an 

 automated ABI 3700 sequencer (Applied Biosystems 

 Inc., Fullerton, CA). 



Because we did not know the identity of the species 

 in the fish fillets, each sequence was submitted to a 

 BLAST (basic local alignment search tool) search on 

 the NCBI GenBank' database, which provides the iden- 

 tification of species by sequences. After we were confi- 

 dent that the identities of the species in the fillets were 

 limited to the genus Perca, the sequences were aligned 

 sequentially with cytochrome b sequence data from all 

 three species of Perca (e.g., P. flavescens, P. fluviatilis, 

 and P. schrenkii), and two outgroup percid taxa (walleye 

 [Sander vitreus] and ruffe [Gymnocephalus cernuus]) by 

 using the computer program CLUSTALX22 (Thompson 

 et al., 1997). A tree of the relationships among the se- 

 quences was constructed by using maximum likelihood 

 estimates of sequence divergences with a neighbor- 

 joining network (Saitou and Nei, 1987) as implemented 

 in the computer program PHYLIP (PHYLogeny Infer- 

 ence Package; Felsenstein^). 



Results and discussion 



Our extraction procedure yielded DNA usable for PCR 

 and subsequent sequencing, although the extractions 

 were degraded and of relatively low molecular weight. 

 Thirteen complete cytochrome b sequences represent- 

 ing all three species of Perca were found documented 

 on GenBank^ (Table 1). Yellow perch is one of three 

 described species of this genus, for which a substantial 

 fishery exists in the Great Lakes, although fish stocks 

 have declined at some localities.^ The natural distribu- 

 tion of yellow perch extends from Nova Scotia south 

 along the Atlantic coast of North America to South Caro- 

 lina, and west to Montana (Scott and Grossman, 1973; 

 Craig, 2000). The Eurasian perch is very similar mor- 

 phologically to P. flavescens, is found throughout most of 



Gibson, T., D. Higgins, J. Thompson, and F. Jeanmougin. 

 2006. ClustalX. Plate-forme de bio-informatique (Bio- 

 informatic platform), LG.B.M.C, 1 rue Laurent Fries, 67404 

 lUkirch, Cedex, France. Website: http://bips.u-strasbg.fr/fr/ 

 Documentation/ClustalX (accessed 14 February 2006). 



' Felsenstein, J. 1995. Department of Genome Sciences, 

 University of Washington, Box 357730, Seattle, Washing- 

 ton, USA 98195-7730. Website: http://evolution. genetics. 

 washington.edu/phylip/general.htmi (accessed 20 February 

 2006). 



' Johnson, T. B. 2006. Personal common. Ontario Ministry 

 of Natural Resources, Glenora Fisheries Station, R.R. #4, 

 21 Hatchery Lane, Picton, Ontario, Canada KOK 2T0. 



100 



100 



r Perca fluviatilis 2 



' P fluviatilis 6 



P fluviatilis 5 

 - P fluviatilis 4' 

 ggi P fluviatilis 1 

 ' P fluviatilis 3' 



C Perca schrenkii 1 

 P schrenkii 2 



98 



I- P schrenkii 3 

 Perca flavescens 2"' 



Eurasian 

 perch 



98 



100 



P flavescens 5 

 P flavescens 3 

 •— P flavescens 1 

 Gymnocephalus cernuus 

 Sander vitreus 



001 



Balkhash 

 perch 



Yellow 

 perch 



Ruffe 

 Walleye 



Figure 1 



Phylogenetic relationships among Perca and outgroup 

 mtDNA haplotypes as inferred from cytochrome b 

 sequence divergences. Taxon labels are consistent with 

 those in Table 1. * = sequence match to an individual 

 fried fish fillet sample sequenced in this study. Numbers 

 at nodes indicate percent support for the relationships 

 from 1000 bootstrap pseudoreplications. 



northern Europe and Asia (Craig, 2000; Maitland, 2000), 

 and also represents a commercially important species. 

 A third species, P. schrenkii Kessler, is restricted to the 

 eastern portion of Kazakhistan and does not contribute 

 to the world market. 



Each species of Perca is represented by unique nu- 

 cleotide cytochrome b sequences and has marked diver- 

 gences (Fig. 1). Three of our five samples matched one 

 of the cytochrome b sequences of the yellow perch P. 

 flavescens (designated as flavescens 2), and two matched 

 sequences of the Eurasian perch P. fluviatilis (denoted 

 as fluviatilis 3 and fluviatilis 4). It is noteworthy that 

 large genetic divergences separate the mtDNA cyto- 

 chrome b sequences of P. flavescens and P. fluviatilis. 

 These two species differ at 130 sites (11.4%; Song et 

 al., 1998; Sloss et al., 2004), and intraspecific variation 

 for each is an order of magnitude less (Fig. 1; also see 

 Billington, 1993). Thus, it is unlikely that our identifi- 

 cations were in error. 



Although one would expect that most of the perch 

 fillets at local markets are caught locally, the supply of 

 and demand for North American and Eurasian perch 

 determines which species is the most economical to 

 serve. Importers and exporters trade fish from both 

 sides of the ocean and the price fluctuates seasonally for 

 both species. Winter is the low supply period for both. 



