KASP genotyping assay and statistical analysis

A total of 110 DNA samples, including 20 and 30 individuals in the bulks resistant and susceptible to isolate Zz26, respectively, and also 60 Malus germplasm accessions were genotyped using the KASP assay (LGC Genomics, Beverly, MA, USA). The primers were designed based on the 200-bp sequence flanking the SNPs (Table S2). The DNA of ‘Jonathan’ and ‘Golden Delicious’ was used as control. Then the genotypes were validated by Sanger sequencing. Data were analyzed using the “Endpoint Genotyping” method of the Light Cycler 480 Software (release 1.5.0).

The segregation biases of markers between resistant and susceptible bulks were tested by Chi-square test. Chi-square test was run twice, the first, allele A was supposed to be completely/partially dominant on B, and the next, allele B completely/partially dominant on A. The scenario which Chi-square value was statistically significant was accepted.