DEVELOPMENT OF MOLECULLAR MARKERS AND GENETIC DIVERSITY OF SAN PEDRO MÁRTIR TROUT Oncorhynchus mykiss nelsoni

Rigoberto Delgado-Vega*, Fabiola Lafarga-de La Cruz, Francisco J. García-de León, Carmen G. Paniagua-Chávez and Miguel Ángel del Río-Portilla.
Departamento de Acuicultura. Centro de Investigación Científica y de Educación Superior de Ensenada (CICESE). Carretera Ensenada-Tijuana No. 3918, Zona Playitas, C.P. 22860, Ensenada, B.C. México
rdelgado@cicese.edu.mx
 

 

In Mexico, there are at least six species of native trout, one of these species is the San Pedro Mártir trout, Oncorhynchus mykiss nelsoni (Evermann, 1908), which is a subspecies of rainbow trout. San Pedro Martir trout lives in the biosphere reserve named Sierra de San Pedro Mártir in Baja California. This organism has characteristics that give aquaculture potential, for example, it can survive in wide temperature range, reaches maturity during the first year of life and also has no migratory behavior. This study can be useful in the conservation of San Pedro Mártir trout, helping to understand the structure and genetic variability of wild populations. Furthermore, these markers could also be useful in selective breeding programs, parentage analysis and to estimates inbreeding. The aim of this study was 1) generate specific microsatellite and single nucleotide polymorphism (SNP) markers for this species using next-generation sequencing and 2) evaluate the genetic diversity of San Pedro Mártir trout.

Eight organisms of San Pedro Mártir trout were collected by electrofishing, and total DNA was extracted from muscle and fin using E.Z.N.A DNA kit. Genomic DNA Isolation Kit, sequencing was conducted using MiSeq platform (Illumina). Assembling was carried out with the CLC Genomics Workbench V. 6.5 program. The microsatellite search was conducted with QDD software. Microsatellites were selected considering the depth of the contig mapping. The SNP discovery was carry out with CLC Genomics Workbench program V. 6.5. Frequency of variation was set at minimum of 12.5% and a depth minimum of 10 reads on the polymorphism site. From the sequencing, a total of 4 688 266 reads were obtained and after assembly, a total of 126 123 contigs were formed. From these contigs, a total of 10 100 contained microsatellite sequences of which it was possible to design primers for 6 842 potential microsatellites. Most of the microsatellites were dinucleotide type (94.02%), followed by tri (4.65%), tetras (1.21%) and pentanucleotide (0.12%). Seventeen tetranucleotide microsatellites complied with the settings for amplifications. These microsatellites were amplified in 40 individuals. In contrast, a total of 99 885 nucleotide variants were found, of which 6 697 were deletion, 6 022 insertions, 5 643 multi nucleotide polymorphism and 81 523 SNPs. The most common type of polymorphism in SNPs was T/C and the least common was C/G. A total of 71 SNPs were selected and the corresponding primers were designed for amplification. In this paper we show and discuss the final results of the genetic diversity of San Pedro Mártir trout using the molecular markers obtained from high throughput sequencing.