GBS of Phytophthora spp.
GBS libraries were prepared from Phytophthora isolates according to the method described by Poland et al. (2012), with some modifications. Approximately 100 ng DNA of each sample was digested with 1 l PstI FastDigest, and 1 l HpaII FastDigest. The PstI adapter containing the barcodes and the HpaII Y-shaped adapter were ligated to the restriction sites with T4 ligase. The ligate was cleaned up using Sera-Mag SpeedBeads. In total, 10 l DNA was used in a PCR mix containing Taq 2x Master Mix (NEB) and 0.75 M each of primer IlluminaF_PE and IlluminaR_PE in a total volume of 50 L. The fragments were PCR-amplified and subsequently 25 l of each sample were cleaned using Sera-Mag SpeedBeads and eluted in 30 l 0.1x TE. Library quality was assessed by quantifying the DNA with a Quantus Fluorometer and by visualizing the fragment length profile using a QIAxcel Advanced System (Qiagen). In total, 96 samples with 96 unique barcodes were pooled for one lane of Illumina HiSeq3000 sequencing. The libraries were paired-end sequenced on an Illumina HiSeq3000 instrument (2x150 bp) at the Oklahoma Medical Research Foundation Genomics Facility (Oklahoma City, OK, USA). All read data were demultiplexed.