title: ITS1 and ITS2 of Erysimum popovii
design: We constructed two libraries, one for ITS1 amplicons and one for ITS2 amplicons. The libraries were prepared using the Nextera XT DNA Sample Preparation Kit. Then, the DNA was amplified via a nine-cycle PCR. The total volume reaction was 25 l with the following composition: 5 L 10 buffer at 1.0 mM (New England BioLabs), 0.1 mM each dNTP, 0.2 M each Nextera primer, 0.02 U Taq high fidelity DNA-polymerase (Q5, NEB), and 5 Q5 High GC Enhancer (NEB). PCR thermocycling conditions were 98 C during 5 s, 55 C for 10 s, and 72 C for 10 s. After that, we purified both libraries using the GenElute PCR Clean-Up Kit (Sigma) to remove short library fragments. We generated equal volumes of the libraries to prepare equimolar libraries for sequencing, and the final concentration of each library was quantified using the Infinite M200 PRO NanoQuant spectrophotometer (TECAN, Mnnedorf, Switzerland). ITS1 and ITS2 library sequencing was carried out in an Illumina MiSeq platform (Illumina, USA) using a paired-end 150 bp sequence read run.