"UCE target enrichment of Cyprinella_venusta_LSUMZ14299: fin/muscle tissue"
We input 100-1000 ng sheared DNA to a modified genomic DNA library preparation protocol (v1.10; http://ultraconserved.org) that incorporated "with-bead" cleanup steps (Fisher et al. 2011) using a generic SPRI substitute (hereafter SPRI; Rohland and Reich 2012). Following adapter ligation, 50% of the resulting library volume (~15 _L; 50-500 ng) was PCR amplified using a reaction mix of 25 _L HiFi HotStart polymerase (Kapa Biosystems), 5 _L of Illumina TruSeq primer mix (5 _M each), and 5 _L ddH20 using the following thermal protocol: 98ĄC for 45s; 10-16 cycles of 98ĄC for 15s, 60ĄC for 30s, 72ĄC for 60s; and a final extension of 72ĄC for 5m. Resulting reactions were purified using 1X SPRI, and we re-hydrated libraries in 33 _L ddH2O. We quantified 2 _L of each library using a Qubit fluorometer. Groups of eight libraries at equimolar ratios were combined into enrichment pools having a final concentration of 147 ng/_L. Using a custom target enrichment bait kit from MYcroarray Inc. (Faircloth et al. 2012; McGee et al. 2016) a standard UCE enrichment protocol was followed (v1.4; http://ultraconserved.org). Based on the size-adjusted concentrations estimated by qPCR, we created an equimolar pool of libraries at 10 nM concentration, and we sequenced 9-10 pmol of each pool-of-pooled libraries using either (Supplemental Table 2) paired-end, 250 bp sequencing on an Illumina MiSeq (v2; UCLA Genotyping Core Facility) or paired-end 100 bp sequencing on an Illumina HiSeq (v3; UCLA Neuroscience Genomics Core).