2007 Proc. Annu. Conf. SEAFWA Genetic Marker-assisted Restoration of the Presumptive Native Walleye Fishery in the New River, Virginia and West Virginia George C. Palmer, Virginia Department of Game and Inland Fisheries, 1796 Highway 16, Marion, VA 24354, USA Joe Williams, Virginia Department of Game and Inland Fisheries, 2206 South Main Street, Blacksburg, VA 24060, USA Mark Scott, West Virginia Division of Natural Resources, 2006 Robert C. Byrd Drive, Beckley, WV 25801, USA Kathy Finne, Department of Fisheries and Wildlife Sciences, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0321 Nathan Johnson, Department of Fisheries and Wildlife Sciences, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0321 Daniel Dutton, Department of Fisheries and Wildlife Sciences, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0321 Brian R. Murphy, Department of Fisheries and Wildlife Sciences, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0321 Eric M. Hallerman, Department of Fisheries and Wildlife Sciences, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0321 Abstract: Te increasing importance of the walleye (Sander vitreus) fshery in the New River, Virginia, and recent research fndings showing persistence of a presumptive native stock motivated a seven-year program of hatchery-based restoration of the native fshery. Candidate spawners were collected from spawning areas, and DNA from fn clips was genotyped at two microsatellite loci. Candidates exhibiting alleles at the Svi17 and Svi33 loci that characterize the presumptive native stock were spawned. Teir young were reared at one of four fsh hatcheries in Virginia and West Virginia. Approxi- mately 600,000 fry and 800,000 fngerlings were stocked in riverine sections of the New River in Virginia and West Virginia. Since stocking began, ages 0–3 walleye have become much more abundant at the upper New River spawning areas. Catch rates in spring electrofshing samples have increased from 3 to 17 fsh per hour in Virginia and from 1.2 to 26.6 fsh per hour in West Virginia. Walleye catch per net-night from fall gill net sets in Claytor Lake, Virginia increased from 0.2 in 2001 to 3.1 in 2006. Comparison of data from creel surveys in 2002 and 2007 showed increasing angler efort di- rected toward walleye from 10% to 30% and increasing catch. Native walleye allele frequencies increased from 16% and 14% at the two marker loci in the 1997 to 1999 genetic surveys to 46% and 58% in the 2004 to 2006 surveys. Key words: walleye, Sander vitreus, microsatellite DNA, marker-assisted selection, supplemental stocking. Proc. Annu. Conf. Southeast. Assoc. Fish and Wildlife Agencies 61:17–22 Walleye (Sander vitreus) is a highly valued sportfsh that inhab- its the New River in southwestern Virginia and West Virginia. Te New River is located on the eastern edge of the native range, and it is uncertain whether walleye is native to the drainage (Jenkins and Burkhead 1994). Hackney and Holbrook (1978) believed walleye to be native to the New River and part of a southern stock found throughout the Mississippi drainage. In the past decade, ten 5- to 7-kg walleyes were collected by anglers or the Virginia Depart- ment of Game and Inland Fisheries (VDGIF) in the upper New River above Claytor Lake. Walleyes have been stocked outside of their native range to ar- eas throughout the United States (Hackney and Holbrook 1978). Introductions from diferent geographic origins resulted in many areas containing mixtures of native and introduced stocks (Mur- phy et al. 1983, Fox 1993, Jennings et al. 1996, Eldridge et al. 2002, White et al. 2005, Palmer et al. 2006). Walleyes of diferent geo- graphic origins were mixed in Claytor Lake and the upper New River, Virginia, as a direct result of planting in 27 stocking events from 1939 to 1996 (Jenkins and Burkhead 1994, Murphy et al. 1983, Palmer 1999). Genotype frequencies for isozyme markers indicated mixing of walleye stocks in Claytor Lake, showing that at least some of these plantings were successful (Murphy et al. 1983). All stocking was suspended in 1997. Movements of radio-tagged fsh showed three spawning sites (Allisonia, Fosters Falls, and Buck Dam, Fig. 1) and suggested that lake- and river-dwelling individu- als to some degree spawned in spatially distinct areas (Palmer et al. 2005). A genetic study showed fsh carrying three previously unknown mitochondrial DNA haplotypes and high frequencies of charac- teristic alleles at particular microsatellite DNA loci (Palmer et al. 2006). Mitochondrial haplotype 43-bearing walleye from the up- per New River tended to exhibit particular alleles at two micro- satellite loci. At the Svi17 locus, the 99/99-homozygous genotype was observed in 94% of all haplotype 43-bearing individuals. Tis 17