Whole genome sequencing of NISTCHO
Samples of Clone 31 research cell bank (RCB) cultures were collected within 2 passages of thaw and were prepared using the DNeasy Blood & Tissue Kit (Qiagen, cat. 69504) using a QIACube (Qiagen) automated DNA extraction platform according to the manufacturers protocol. Purified DNA was quantified via Nanodrop (ThermoFisher) or using a Qubit fluorometer (ThermoFisher). NEBNext Ultra DNA Library Prep Kit for Illumina, clustering, and sequencing reagents was used throughout the process following the manufacturers recommendations. Briefly, the genomic DNA was fragmented by acoustic shearing with a Covaris S220 instrument. Fragmented DNA was cleaned up and end repaired. Adapters were ligated after adenylation of the 3ends followed by enrichment by limited cycle PCR. DNA libraries were validated using a High Sensitivity D1000 ScreenTape on the Agilent TapeStation (Agilent Technologies, Palo Alto, CA, USA), and were quantified using Qubit 2.0 Fluorometer. The DNA libraries were also quantified by real time PCR (Applied Biosystems, Carlsbad, CA, USA). The sequencing library was clustered onto a flowcell. After clustering, the flowcell was loaded onto the Illumina HiSeq 4000 instrument according to manufacturers instructions. The samples were sequenced using a 2x150bp Paired End (PE) configuration.