Performance comparison of gel and capillary electrophoresis-based microsatellite genotyping strategies in a population research and kinship testing framework

Ever since their discovery in 1981 [1], the usefulness of microsatellites, due to their hypervariability and ubiquitous occurrence, has been astounding to geneticists [2]. Microsatellites are short tandem repeats of almost anything from 1 to around 6 bp [2]. They are highly informative, codominant, and transferable among phylogenetically-related species [3]. As such, microsatellites are powerful tools commonly used in population research to infer genetic diversity, genetic structure, and mating systems [4,5,6]. Microsatellites have also found their application in linkage-disequilibrium, in which associations between markers and traits are searched, and hitchhiking mapping, in which genome-wide surveys are used to identify regions showing positive selection [7, 8]. However, to obtain reliable information, microsatellite genotyping should be performed with extreme accuracy since high error rates could inject bias into the downstream analysis and, therefore, alter ecological and evolutionary conclusions [9, 10].

Besides allelic dropouts and null alleles, the most common errors during microsatellite genotyping include contaminant DNA, incorrect data entry, and scoring errors [11]. Although most of these errors can be detected if the genotypification is repeated, the proportion of mis-scored alleles, which can be up to 80%, will depend on the researchers’ genotyping method [12]. The gold standard for microsatellite genotyping is capillary electrophoresis (CE), a technology that accurately scores the alleles owing to its technology that implements automated allele-call programs [13]. However, because this method requires sophisticated instruments, it is usually impractical for many laboratories in countries having no access to cutting-edge technologies [14, 15]. Nonetheless, more costly-effective methods such as polyacrylamide gel electrophoresis (PAGE) have been remarkably resilient in competition against sequencing techniques [16], and are still being utilized by researchers, especially in population research [17,18,19]; though the accuracy of their results compared to those obtained using high-throughput (HTP) methods is still arguable [12, 20,21,22].

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Here, we present a direct comparison between PAGE and CE methods by applying them in the genotypification of five microsatellite loci in an economically important fish population. To the best of our knowledge, this study is the first of its kind that compares not only allele frequency data but also the different genetic diversity parameters analyzed in population research. Moreover, this study evaluates the performance of the PAGE method by comparing the results of relatedness analysis to those obtained using the CE method.

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