Three mapping populations were used in this study: (1) F2 populations of SP2271 × SP2272 and (2) BC1 and (3) F2 populations of SP2271 × SP2300. The populations were phenotyped by agroinfiltration of RXLR effectors. A cork borer was used for sampling, and the leaf discs from the responsive and non-responsive progenies were pooled. The Genomic DNA was isolated using the Qiagen DNeasy plant kit (Qiagen, 69104). RenSeq libraries were then prepared, as described previously24 (link). The libraries were sequenced (Illumina 2 × 250-bp reads) by Novogene (Beijing, China). The SNP filtering and calling steps were described previously26 (link).
To design molecular markers, the 10× PCR-free resequencing reads were mapped to the SP2271 S. americanum reference genome. Then SCAR markers that linked with the BSA-RenSeq signals were designed; amplicons should only be present in the non-responsive allele. The SCAR markers were first tested on the parental lines and the verified markers were then used on genomic DNA from individual non-responsive plants. GoTaq G2 DNA polymerase (Promega, 0000066542) was used for genotyping.
To design molecular markers, the 10× PCR-free resequencing reads were mapped to the SP2271 S. americanum reference genome. Then SCAR markers that linked with the BSA-RenSeq signals were designed; amplicons should only be present in the non-responsive allele. The SCAR markers were first tested on the parental lines and the verified markers were then used on genomic DNA from individual non-responsive plants. GoTaq G2 DNA polymerase (Promega, 0000066542) was used for genotyping.
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