a Concentration of each primer was 10 µM. PCR-based
sexing (Rds1 and Rds2) was carried out in a total volume of 16 µl, while
HRM (Rds3) was carried out in 15 µl reaction mixture.
b PCR-based sexing of Rds3 performed best under the
conditions shown here. Binding of the Y-primer was SNP-specific, but
band intensities on agarose gel were often insufficient (i.e. neither
the X/Y universal nor the Y-specific products were detectable in many
cases), therefore we used the HRM method instead.
Primer: primer names follow the logic shown in Figure S1, where F means
universal forward, R means universal reverse and Y-F and Y-R means
Y-specific forward and reverse primers, respectively.
PCR ID: PCR programs are described in Table S3.
Y-SNPs are denoted with bold underlined letters and artificial
mismatches (Liu et al., 2012) are bold.