Benefits and limitations of a new genome-based PCR-RFLP genotyping assay
(GB-RFLP): a SNP-based detection method for identification of species in
extremely young adaptive radiations
Abstract
High-throughput DNA sequencing technologies make it possible now to
sequence entire genomes relatively easily. Complete genomic information
obtained by whole genome resequencing (WGS) can aid in identifying and
delineating species even if they are extremely young, cryptic or
morphologically difficult to discern and closely related. Yet for
taxonomic or conservation biology purposes WGS can remain
cost-prohibitive, too time-consuming, and often constitute a “data
overkill”. Rapid and reliable identification of species (and
populations) that is also cost-effective is possible based on
species-specific markers that can be discovered by WGS. Based on WGS
data we designed a PCR restriction fragment length polymorphism
(PCR-RFLP) assay for 19 Neotropical Midas cichlid populations
(Amphilophus cf. citrinellus), that includes all 13 described species of
this species complex. Our work illustrates that identification of
species and populations (i.e., fish from different lakes) can be greatly
improved by designing genetic markers using available “high
resolution” genomic information. Yet, our work also shows that even in
the best-case scenario, when whole-genome resequencing information is
available, unequivocal assignments remain challenging when species or
populations diverged very recently, or gene flow persists. In summary,
we provide a comprehensive workflow on how to design RFPL markers based
on genome re-sequencing data, how to test and evaluate their
reliability, and discuss the benefits and pitfalls of our approach.