Qubit 2 system

Pure DNA samples used in this study were obtained via a modified phenol-chloroform extraction protocol [46 (link)]. DNA samples were quantified using the Quant-iT^™ dsDNA High Sensitivity assay on the Invitrogen^™ Qubit^™ 2 system (Thermo Fisher Scientific). Serial dilutions of Phytophthora ramorum and P. cinamommi were made to determine the LOD of the assay, as explained below.
Crude plant extracts were made from healthy Umbellularia californica leaves. Briefly, 0.5 g of leaf tissue was added to 5 mL to GEB2 buffer within a netted bag and ground. A volume of 1 µL of this crude plant extract was added to purified DNA to determine the LOD of the assay with and without a crude plant extract, as described below.

DNA was salted out (Miller et al. 1987 ) from EDTA blood to produce high-quality DNA. DNA concentration was initially measured with the Nanodrop 2000 spectrophotometer (Thermofisher, USA) and diluted to 20–30 ng/µl. Before library preparation for next-generation sequencing, diluted DNA was quantified again with the Qubit 2 system (Thermofisher, USA) using the Qubit dsDNA broad range assay kit and the Qubit dsDNA high sensitivity assay kit. Subsequently, paired-end libraries were prepared with the TruSeq DNA PCR-free sample preparation kit (Illumina, USA). Generated libraries were quantified through qPCR with the Kapa Library Quantification Kit (Kapabiosystems, USA) and distribution of fragment sizes in the libraries was checked with the BioAnalyzer 2100 (Agilent Genomics, USA).
Whole genome sequencing was performed on the Illumina HiSeq2500 (Illumina, USA). Paired-end libraries (2 × 126 bp reads) were sequenced with a mean coverage of ten times (n = 48 animals) and four times (n = 10 animals), respectively.

Genomic DNA was extracted from tissue samples and antler drill cores with the Instant Virus RNA Kit (Analytik Jena, Germany). Antler drill cores (0.1 to 0.3 g) were treated in a bead mill (MM200, Retsch, Germany) at a frequency of 25 Hz for 2 min. Tissue samples were suspended in 450 μl of lysis buffer and subsequently treated in the same way as the antler drill cores. All following steps were performed according to the manufacturer’s instructions. The extracted DNA was eluted with 60 μl of RNAse-free water.
DNA concentration was measured photometrically with the Nanodrop 2000 spectrophotometer (Thermofisher, USA) and the Qubit 2 system (Qubit dsDNA br assay kit and Qubit dsDNA hs assay kit, Thermofisher, USA).