A high molecular weight (HMW) DNA extraction was carried out for HM078 using the Monarch HMW DNA Extraction Kit for Cells & Blood (New England Biolabs, Ipswich, MA) following the kit’s protocol for cells. The Short Read Eliminator Kit (Circulomics Inc., Baltimore, MD, USA) was used to deplete DNA fragments below 25 kbp. The resulting HMW DNA was quantified with a Qubit 3.0 Fluorometer (Thermo Fisher Scientific, Waltham, MA, USA), and approximately 200 ng was run on a 0.8% agarose gel along with the NEB Quick-Load 1 kbp Extend DNA ladder (New England Biolabs, Ipswich, MA). Pulse-field capillary electrophoresis (FEMTO-Pulse, Agilent Technologies Inc., Santa Clara, CA, USA) was utilized to assess the relative quantity of HMW DNA at various fragment lengths. HMW DNA was stored at 4°C until library preparation.
PacBio long-read DNA sequencing was conducted at the University of Minnesota, Twin Cities, Genomics Center. Prior to library preparation, HMW DNA was purified with AMPure PB beads (Pacific Biosciences, Menlo Park, CA, USA). The PacBio SMRTbell prep kit 3.0 (Pacific Biosciences, Menlo Park, CA, USA) was used to for library preparation following the manufacturer’s specifications, and long-read sequencing was performed on the Sequel II system with two 8M SMRT Cells running for a 30-hour period (Pacific Biosciences, Menlo Park, CA, USA).
PacBio long-read DNA sequencing was conducted at the University of Minnesota, Twin Cities, Genomics Center. Prior to library preparation, HMW DNA was purified with AMPure PB beads (Pacific Biosciences, Menlo Park, CA, USA). The PacBio SMRTbell prep kit 3.0 (Pacific Biosciences, Menlo Park, CA, USA) was used to for library preparation following the manufacturer’s specifications, and long-read sequencing was performed on the Sequel II system with two 8M SMRT Cells running for a 30-hour period (Pacific Biosciences, Menlo Park, CA, USA).