High‐Resolution Melting Analysis for the Discovery of Novel Single‐Nucleotide Polymorphisms in Rainbow and Cutthroat Trout for Species Identification

Abstract We describe the use of high-resolution melting (HRM), a recent enhancement to traditional DNA melting analyses for the characterization of polymerase chain reaction products, in the identification of 11 novel single-nucleotide polymorphisms (SNPs) for distinguishing between rainbow trout Oncorhynchus mykiss and cutthroat trout O. clarkii. Single-nucleotide polymorphisms provide an important tool for species identification in studies of hybridization and introgression between these two species, but until now the time-consuming and costly nature of DNA sequencing has limited identification of new markers. The ability of HRM to accurately discern nucleotide changes in a DNA sequence make it a cost- and time-effective alternative to traditional sequencing for the detection of novel SNPs.

[1]  W. R. Ardren,et al.  Morphological, Physiological, and Genetic Techniques for Improving Field Identification of Steelhead, Coastal Cutthroat Trout, and Hybrid Smolts , 2009 .

[2]  B. May,et al.  Subspecies‐informative SNP assays for evaluating introgression between native golden trout and introduced rainbow trout , 2009, Molecular ecology resources.

[3]  L. Seeb,et al.  SNP genotyping by the 5'-nuclease reaction: advances in high-throughput genotyping with nonmodel organisms. , 2009, Methods in molecular biology.

[4]  F. Allendorf,et al.  Rainbow trout (Oncorhynchus mykiss) invasion and the spread of hybridization with native westslope cutthroat trout (Oncorhynchus clarkii lewisi) , 2008 .

[5]  Andrew P. Martin,et al.  Hybridization dynamics between Colorado's native cutthroat trout and introduced rainbow trout. , 2008, The Journal of heredity.

[6]  K. Voelkerding,et al.  Quadruplex genotyping of F5, F2, and MTHFR variants in a single closed tube by high-resolution amplicon melting. , 2008, Clinical chemistry.

[7]  M. Campbell,et al.  Incidence of Hybridization between Naturally Sympatric Westslope Cutthroat Trout and Rainbow Trout in the Middle Fork Salmon River Drainage, Idaho , 2007 .

[8]  Robert Palais,et al.  Simultaneous mutation scanning and genotyping by high-resolution DNA melting analysis , 2007, Nature Protocols.

[9]  P. Smouse,et al.  genalex 6: genetic analysis in Excel. Population genetic software for teaching and research , 2006 .

[10]  L. Seeb,et al.  Use of sequence data from rainbow trout and Atlantic salmon for SNP detection in Pacific salmon , 2005, Molecular ecology.

[11]  B. Porfirio,et al.  Sequence diversity within the HA-1 gene as detected by melting temperature assay without oligonucleotide probes , 2005, BMC Medical Genetics.

[12]  F. Allendorf,et al.  Intercrosses and the U.S. Endangered Species Act: Should Hybridized Populations be Included as Westslope Cutthroat Trout? , 2004 .

[13]  Carl T Wittwer,et al.  Sensitivity and specificity of single-nucleotide polymorphism scanning by high-resolution melting analysis. , 2004, Clinical chemistry.

[14]  Elaine Lyon,et al.  Genotyping of single-nucleotide polymorphisms by high-resolution melting of small amplicons. , 2004, Clinical chemistry.

[15]  G. Luikart,et al.  SNPs in ecology, evolution and conservation , 2004 .

[16]  C. Ostberg,et al.  Bi-parentally inherited species-specific markers identify hybridization between rainbow trout and cutthroat trout subspecies , 2003 .

[17]  D. Heath,et al.  Erosion of interspecific reproductive barriers resulting from hatchery supplementation of rainbow trout sympatric with cutthroat trout , 2003, Molecular ecology.

[18]  Carl T Wittwer,et al.  High-resolution genotyping by amplicon melting analysis using LCGreen. , 2003, Clinical chemistry.

[19]  P. Kwok Single nucleotide polymorphisms : methods and protocols , 2002 .

[20]  P. Bentzen,et al.  Molecular Markers Distinguish Coastal Cutthroat Trout from Coastal Rainbow Trout/Steelhead and Their Hybrids , 2002 .

[21]  Joseph R. Tomelleri,et al.  Trout and Salmon of North America , 2002 .

[22]  A. M. Garrett,et al.  Status review of coastal cutthroat trout from Washington, Oregon, and California , 1999 .

[23]  D. Hawkins,et al.  Early survival and development of coastal cutthroat trout (Oncorhynchus clarki clarki), steelhead (Oncorhynchus mykiss), and reciprocal hybrids , 1998 .

[24]  F. Bonhomme,et al.  GENETIX 4.05, logiciel sous Windows TM pour la génétique des populations. , 1996 .

[25]  François Rousset,et al.  GENEPOP (version 1.2): population genetic software for exact tests and ecumenicism , 1995 .

[26]  F. Allendorf,et al.  Conservation and Distribution of Genetic Variation in a Polytypic Species, the Cutthroat Trout , 1988 .

[27]  F. Allendorf,et al.  DEVELOPMENTAL INSTABILITY AND HIGH MERISTIC COUNTS IN INTERSPECIFIC HYBRIDS OF SALMONID FISHES , 1985, Evolution; international journal of organic evolution.

[28]  F. Utter,et al.  Natural hybridization between steelhead trout (Salmo gairdneri) and coastal cutthroat trout (Salmo clarki clarki) in two Puget Sound streams , 1985 .