DNA-seq of homo sapiens: adult male blood
For Hifi long-read sequencing of APR cohort samples, high molecular weight DNA was isolated from 200 L of flash frozen blood using both GenFind V3 and Nanobind DNA extraction kits per manufacturers instructions. For shearing, DNA concentration was first measured using Qubit 3 Fluorometer with a dsDNA HS Assay kit (Thermo Fisher) and samples were adjusted to 30 ng/L in a total volume of 130 L. DNA samples were then loaded on Diagenode Megaruptor 3 hydropore-Syringe at a speed of 29 targeting 15-18 kb fragment length as recommended by the PacBio protocol. The sheared samples were run on 4200 Tapestation system to check the fragment length using Genomic DNA screen tape. SMRTbell libraries were prepared for sequencing using the SMRTbell Prep Kit 3.0. The libraries were attached to Sequel II primer 3.2 and Sequel II DNA polymerase 2.2 and sequenced on PacBio Sequel IIe equipment using SMRT cells 8M and Sequel II Sequencing kit 2.0. For each sample on Sequel 11e, three SMRT cells were used to produce at least 30x coverage. The runs were created to allow adaptive loading, 2h of pre-extension and 30h of movie times