C_americana
I used a double digest restriction site associated DNA sequencing (ddRADseq) protocol to obtain SNPs from across the genome (Peterson et al. 2012). I performed DNA extractions on either fresh or dried tissue in 96-well plates using a modified CTAB chloroform extraction protocol. Next gen sequencing libraries were built with a Biomek NXp liquid handling robot. Samples were randomized within plates, then digested with the enzymes of ApoI HF and Sph! HF simultaneously. I then ligated one of 48 unique adapters to the DNA of each sample, purified with ampure beads, and size selected for fragments of 450-550bp using a BluePippin machine. After size selection, I made 9 pools of 48 samples, each with unique adapters, and performed PCR on each pool using a different primer for each one, given each sample a unique combination of indices. PCR products were normalized based on qPCR, then pooled and sent to BGI for sequencing on two lanes of Illumina HiSeq 4000. Samples here are demultiplexed.