The eastern indigo snake (Drymarchon couperi) is the longest snake in North America. Population declines due to extensive loss of habitat led to federal listing as a threatened species in 1978. Knowledge deficiencies regarding the biology of the eastern indigo snake, including population connectivity and gene flow, are impediments to the development of effective conservation and restoration strategies. We describe primers and polymerase chain reaction (PCR) conditions to amplify 22 tetranucleotide and pentanucleotide nuclear microsatellite loci isolated from the eastern indigo snake. We tested primers using 28 shed skins representing 24 individuals from Fort Stewart, Georgia. Primers yielded an average of 4.6 alleles per locus (range 3-7) and an average observed heterozygosity of 0.63 (range 0.46-0.79). These loci should prove useful for individual identification as well as population level analyses of this federally threatened species.
The leatherback turtle (Dermochelys coriacea) is a globally endangered marine species. Numerous questions regarding life history and demographics that are of conservation interest remain and many of these can be addressed through the use of highly polymorphic nuclear markers. We describe primers and polymerase chain reaction conditions to amplify 19 tetranucleotide microsatellite loci from the leatherback turtle. The primers were tested on samples from 22 females that nested at Archie Carr National Wildlife Refuge, Melbourne Beach, Florida, USA. The primers developed in this study yielded an average of 9.4 alleles per locus (range of 5-19) and an average observed heterozygosity of 0.84 (range 0.36-1.00). These markers should prove useful in supplementing existing markers for individual and population level analyses.
We describe 12 polymorphic tetranucleotide and pentanucleotide loci in the red-cockaded woodpecker (Picoides borealis). An average of 6.25 alleles per locus was identified based on a screening of 21 individuals from the Joseph W. Jones Ecological Center in southwestern Georgia. Observed heterozygosity ranged from 0.048 to 0.952. These markers could be used for both population level studies and individual identification.Keywords Red-cockaded woodpecker Á Picoides borealis Á Microsatellite Á Tetranucleotide Á PentanucleotideThe red-cockaded woodpecker (Picoides borealis) is an endangered species native to mature pine forests of the southern United States. Large-scale habitat removal and alteration have drastically reduced P. borealis from a nearly continuous range to small pockets of suitable habitat (U.S. Fish and Wildlife Service 2003). Genetic studies are essential to examine the effects of population fragmentation, bottlenecks and inbreeding on this species. It is also of interest to study the consequences of translocations and other restoration efforts on the genetic diversity and viability of P. borealis. This paper describes the development of locus-specific primers and PCR conditions for amplification of 11 polymorphic tetranucleotide loci and 1 pentanucleotide locus in P. borealis. We included five additional loci that were monomorphic in our samples, but may be useful outside this study population or for interspecies utilization (Table 1).Microsatellite loci were isolated using the protocol described by Glenn and Schable (2005). Genomic DNA was extracted from a muscle sample taken from a deceased (from natural causes) P. borealis specimen using a Charge Switch Ò magnetic bead isolation kit (Invitrogen). DNA was eluted in 80 ll and digested using RsaI endonuclease (New England Biolabs) at 37°C for 1 h, and 10 ll was separated on a 1% agarose gel for 1 h at 100 V to evaluate digestion. Double-stranded SuperSNX linkers (Glenn and Schable 2005) were prepared and ligated to the digested DNA and ligation success was verified using PCR and agarose gel electrophoresis. All PCR reactions were conducted using an Applied Biosystems 9700 thermocycler. Digested DNA was hybridized to three different mixtures of biotinylated oligonucleotide probes (Integrated DNA
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