BOVID_1896_2kb
Library originated from gDNA extracted from 0.25g aliquots of fresh fecal material isolated from a periparturient Holstein Friesian dairy cow during a follow-up medical examination for metritis. Following extraction using mechanical and chemical lysis and column-based purification and QC, numerous aliquots of gDNA were combined to meet DNA input requirements for each specific DNA fragment size shearing protocol. Each composited gDNA sample was then subjected to mechanical shearing protocols to achieve either ~2, ~5, or ~8 kb fragment sizes via sonication (for 2 and 5 kb) or hydrodynamic centrifugation (for ~8 kb). Following gel-based size selection and bead-based clean-up, each fecal gDNA sample was subjected to probe-based enrichment and library construction utilizing modified and customized procedures adapted from methods of the SureSelectXT V1 platform and custom biotinylated probes. Following these steps, libraries were further subjected to PacBio library creation using PacBio SMRTbell Express Template Preparation Kit without any additional shearing or amplification. Following hairpin adapter ligation, all enriched libraries for circular consensus sequencing were cleaned using nuclease treatment and AMPure PB bead clean-up.