UCE target enrichment of Triepeolus_martini_CMNTO_339_TGGTAGCT+GTTAGACG [Raw reads]
This study used a reduced representation genome sequencing approach, targeting sequence capture of ultraconserved elements (UCEs). Eluted DNA concentrations were measured using a Qubit 3.0 Fluorometer (Thermo Fisher Scientific Inc.) with the Qubit dsDNA High Sensitivity Assay Kit. Extracted and purified DNA was sheared to a target average fragment size of 400600 base pairs using the Q800R3 Sonicator (Qsonica). Fragmented DNA was purified using KAPA Pure Beads (Roche Diagnostics), and dual-indexed Illumina libraries were prepared using the KAPA HyperPrep Kit with the iTru adapter system (Glenn et al., 2019). For each library, a 20 L end repair and A-tailing reaction mixture was prepared and incubated at 20C for 30 min and 65C for 30 min, after which a 35 L adapter ligation reaction mixture was prepared and incubated at 20C for 15 min. Subsequently, a 50 L amplification reaction mixture was prepared for each library, and the adapter-ligated libraries were amplified using the following PCR conditions: 98C for 45 s; 13 or 16 cycles of 98C for 15 s, 60C for 30 s, and 72C for 60 s; and 72C for 5 min. Libraries with similar concentrations were pooled in sets of ten, and the volumes were adjusted to ensure approximately 100 ng of DNA per library. The volume of each pool was subsequently adjusted to reach a target concentration of 72 ng/L. The two-day enrichment process was performed using the myBaits UCE Hymenoptera 2.5Kv2AB kit (Daicel Arbor Biosciences), with the first day following the procedure described in the myBaits manual (v5.03) (with a 24-hour incubated reaction at 65C) and the second following Faircloths post-enrichment limited-cycle PCR protocol (v2.4) (available at www.ultraconserved.org), which is based on Blumenstiel et al. (2010). Post-PCR DNA concentrations were measured using a Qubit 2.0 Fluorometer. The UCE-enriched libraries were sent to Gnome Qubec Inc. for next-generation sequencing on the NovaSeq X Series sequencing platform (Illumina, Inc.) and demultiplexing.