title: PacBio RS II sequencing; Cane toad whole genome sequencing - PacBio library 1
library_construction_protocol: High-molecular weight genomic DNA (gDNA) was extracted from cane toad liver tissue using the genomic-tip 100/G kit (Qiagen, Hilden, Germany). This was performed with supplemental RNase (Astral Scientific, Taren Point, Australia) and proteinase K (NEB, Ipswich, MA, USA) treatment, as per the manufacturer's instructions. Isolated genomic DNA was further purified using AMPure XP beads (Beckman Coulter, Brea, CA, USA) to eliminate sequencing inhibitors. DNA quantity was assessed using the Quanti-iT PicoGreen dsDNA kit (Thermo Fisher Scientific, Waltham, MA, USA), DNA purity using a Nanodrop spectrophotometer (Thermo Fisher Scientific), and integrity using pulse-field gel electrophoresis. SMRTbell libraries were prepared from gDNA using the 10-20kb Genomic preparation kit (Pacific Biosciences). To increase subread length, 15-50 kb BluePippin size selection (Sage Science, Beverly, MA, USA) was performed.