title: RAD-seq of Euhadra sp.: adult foot
design: RAD-seq was used to generate SNP markers for 16 individuals representing four species. The samples included two sinistral E. quaesita populations (n=6) that are largely parapatric with two dextral E. aomoriensis populations (n=6) in East Iwate and South Yamagata, Japan. For comparison, one population of dextral E. senkenbergiana (n=3) and one individual of sinistral E. decorata were used. The 16 individuals were initially prepared as two separate libraries, then proportionally pooled together into a single library to achieve more uniform coverage across individuals. The raw reads were cleaned and processed into putative RAD loci using STACKS 1.8 (Catchen et al. 2013). Reads were trimmed to 45 bp and then quality checked using a 10% sliding window; if the average phred score dropped below 20 in any window the read was discarded. Reads were also discarded if the barcode region, or SbfI restriction-site contained errors, or if adapter sequence was detected in the read. In the case of three samples it was clear that a substantial number of reads had been discarded due to a single error in the first base of the barcode; therefore these sequences were recovered.