title: WES of HL60: Pyronin Y high and low vs control
design: For whole exome sequencing, genomic DNA was isolated using QIAamp DNA Mini kit (Qiagen, Hilden, Germany). The quantity and quality of the isolated DNA was assessed using NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA USA). Library preparation and sequencing were performed by Novogene Co., Ltd (Beijing, China). Briefly, sequencing libraries were generated using an Agilent SureSelect Human All Exon Kit V6 (Agilent Technologies, Santa Clara, CA, USA). Then, 150-bp paired-end sequencing was performed on the Illumina NovaSeq platform (Illumina, San Diego, CA, USA) using the constructed DNA libraries for WES. Untreated cells were used as controls. Raw sequencing reads (FASTQ files) were first assessed for quality using FastQC (v0.12.1). Adapter sequences and low-quality bases were trimmed using Cutadapt (v3.5) with removal of Illumina adapter sequence (AGATCGGAAGAGC), a minimum read length of 30 bp, quality trimming threshold of Q20, and removal of terminal ambiguous bases. Trimmed reads were aligned to the human reference genome (GRCh38 primary assembly) using BWA-MEM (v0.7.17). PCR duplicates were marked using GATK MarkDuplicates (v4.6.2). For Small Variant Calling, Single nucleotide variants (SNVs) and small insertions/deletions (indels) were identified using GATK (v4.6.2). Variant calling was performed using HaplotypeCaller in GVCF mode for each sample. Individual GVCFs were combined using CombineGVCFs, followed by joint genotyping with GenotypeGVCFs to generate a multi-sample VCF file. Structural variants were identified using Manta (v1.6.0). Deduplicated BAM files were provided as input, and structural variant discovery was performed using the GRCh38 reference genome.
library_name: Control_rep1