Validation of SSR-tetranucleotide multiplex panel for kinship evaluation in tilapia breeding programs

Abstract

This study aimed to develop PCR assays, reaction combinations, and validation of tetranucleotide SSR loci
for tilapia, to minimize the impact of erroneous allele inferences on genotype determination of this marker. Microsatellites
containing tetranucleotide repeats were obtained from tilapia genome, version 2.1, avoiding loci in the same linking group.
Primers were designed for different fragment sizes, and fluorescence added to each locus. A total of 10 loci were amplified,
separately and in combination, and loaded into a single capillary sequencer panel. Alleles were amplified without stutters and
easily interpreted. PCR amplifications of DNA repeatedly extracted from samples, and genotyping at different PCR rounds
were performed to infer allele signaling errors. The panel obtained in this study is currently used in kinship analyses and
pedigree corrections in a tilapia breeding program.