3.2 GB-RFLP marker analysis for the assignment of
sympatric crater lake species
Next, we designed markers to distinguish the sympatric species of crater
lakes Apoyo (A. astorquii , A. chancho , A.
flaveolus , A. globosus , A. supercilius and A.
zaliosus ) and Xiloá (A. amarillo , A. sagittae , A.
viridis and A. xiloaensis ). Although all sympatric species
clearly form separate genetic clusters, the number of shared alleles is
extensive due to ongoing gene flow and/or recent divergence (Kautt et
al., 2016, 2018). Indeed, there are no alternatively fixed differences,
but we show that there are some nearly fixed ones based on which we
should reach 90 to 100% accuracy — however it is possible that these
predictions might be limited because our genomic samples do not
accurately enough reflect the actual population frequencies.
Overall, the quality of the species-specific RFLP markers (Fig. S4) was
much lower than the lake specific markers (Fig. S3). The percentage of
correctly assigned species ranged from 50% (equal to a random
assignment) to 100%. Only two markers achieved values above 90%
(A. chancho , Fig. S4F and A. viridis , Fig. S4O). In
contrast to the lake-specific markers, species specific-markers showed a
much lower accuracy in the GB-RFLP assay compared to the bootstrapping
dataset with 12/16 samples having >20% less correctly
assigned samples than in the bootstrapping dataset (Fig. 3D–F), which
is substantially different from the lake-specific markers (1/18). The
combination of markers did not improve the accuracy.