title: LPS5
design: Genomic DNA was quantified using the Qubit dsDNA High Sensitivity Assay Kit (Thermo Fisher Scientific). Library preparation was performed using the Twist Library Preparation Kit 2.0 (Twist Bioscience) according to the manufacturer&#39;s protocol, with minor optimizations. Briefly, 50 ng of genomic DNA was enzymatically fragmented to achieve an average insert size of approximately 350 base pairs. Fragmentation was followed by end-repair and A-tailing in a single enzymatic step. Universal adapters provided in the kit were ligated to the A-tailed DNA using a high-efficiency ligase enzyme mix. Adapter-ligated fragments were then purified using Alexgen DNA Magbeads (Cat.no.1013) magnetic beads (Alexius Biosciences) at a 1.8 bead-to-sample ratio to remove unligated adapters and small DNA fragments. Following adapter ligation and cleanup, the libraries were amplified by PCR using Twist Unique Dual Index (UDI) primers. PCR was performed for 4 cycles. Final libraries were assessed for fragment size distribution using the Agilent TapeStation D1000 ScreenTape (Cat no. 5067-5582), with expected median sizes around 450 bp, including adapters. Library concentration was determined using Qubit, and libraries were normalized to 2 nM prior to pooling. Pooled libraries were sequenced on an Illumina NovaSeq Xplus platform using a paired-end 150 bp read configuration, targeting a minimum average genome coverage of 1.2 to 1.5x per sample. Raw sequencing data were demultiplexed using bcl2fastq v2.20 (Illumina), and resulting FASTQ files were used for downstream bioinformatic processing.
library_name: LPS5