Express template prep kit v2 and smrtbell cleanup kit v2

PacBio HiFi data were generated from the HG00733 lymphoblastoid cell line as previously described (G. A. Logsdon et al., 2021 (link)) with modifications. Briefly, DNA was extracted from 4.3×10^6 cells using the Monarch HMW DNA Extraction Kit for Cells and Blood (New England Biolabs) with 1400 rpm lysis speed. After UV absorption and fluorometric quantification (Qubit High Sensitivity DNA kit, Thermo Fisher) on the DS-11 FX instrument (Denovix) and evaluation of DNA integrity on FEMTO Pulse (Agilent), 12 μg of DNA was prepared for sequencing using Megaruptor 3 shearing (Diagenode, settings 19/31) and the Express Template Prep Kit v2 and SMRTbell Cleanup Kit v2 (PacBio). The library was size-selected on a PippinHT instrument (Sage Science) using a 15 kbp high-pass cut. Five SMRT Cell 8Ms were run on a Sequel II instrument using Sequel II chemistry C2.0/P2.2 with 30-hour movie times, 2-hour pre-extension, and adaptive loading targets of 0.8–0.85 (PacBio). Circular consensus calling was performed with CCS version 6.0.0 (SMRT Link v.10.1) and reads with estimated quality scores ≥Q20 were selected for downstream analysis.

PacBio HiFi data were generated from the HG00733 lymphoblastoid cell line as previously described (Logsdon et al. 2021 (link)) with modifications. Briefly, DNA was extracted from 4.3 × 10⁶ cells using the Monarch HMW DNA Extraction Kit for Cells and Blood (New England Biolabs) with 1400 rpm lysis speed. After UV absorption and fluorometric quantification (Qubit High Sensitivity DNA kit, Thermo Fisher Scientific) on the DS-11 FX instrument (Denovix) and evaluation of DNA integrity on FEMTO Pulse (Agilent), 12 μg of DNA was prepared for sequencing using Megaruptor 3 shearing (Diagenode, settings 19/31) and the Express Template Prep Kit v2 and SMRTbell Cleanup Kit v2 (PacBio). The library was size-selected on a PippinHT instrument (Sage Science) using a 15 kbp high-pass cut. Five SMRT Cell 8Ms were run on a Sequel II instrument using Sequel II chemistry C2.0/P2.2 with 30-h movie times, 2-h pre-extension, and adaptive loading targets of 0.8–0.85 (PacBio). Circular consensus calling was performed with CCS version 6.0.0 (SMRT Link v.10.1) and reads with estimated quality scores ≥Q20 were selected for downstream analysis.