GSM8527696: KHYG-1 NKp44 KO + collection of mutant cells, round 2.; Homo sapiens; OTHER
genomic DNA was isolated from the cryopreserved cell pellet using the DNeasy Blood and Tissue Kit (Qiagen) and the genomic DNA concentration was determined using a NanoDrop spectrophotometer. Library preparation requires a two-step PCR amplification, purification and quantification according to Webster et al. 2019. In a first PCR step, the sgRNA sequences are amplified with the PCR primers U6-F1 and Tracr-R1. A second PCR was performed to attach Illumina-compatible sequencing adapters for the NextSeq2000 sequencer (genomic platform at CIML). After purification on a 2% size-selective E-gel, the secondary PCR product was quantified using the Qubit dsDNA HS Assay Kit (Thermo Fisher) and qualified on the bioanalyzer (Agilent) using the High sensitivity DNA Assay (Agilent).