高分子量DNA的完整性对于准确的长读长测序和全面的癌症基因组学至关重要

Accurate genomic characterization in oncology and cancer research depends on the quality and integrity of input DNA. Conventional genomic DNA (gDNA) extraction kits often produce short or degraded fragments due to excessive mechanical or chemical disruption during lysis, multiple manual transfer steps, and prolonged elution conditions, which limit the utility of extracted DNA for long-read sequencing and structural variant analysis. Short-read sequencing (50-600 bp) further compounds these limitations by providing incomplete assemblies and reduced ability to resolve large-scale rearrangements. Long-read technologies such as Oxford Nanopore and PacBio overcome these issues but require intact, high molecular weight (HMW) DNA fragments (>10 kb). To address these challenges, genomic DNA was extracted from whole blood, cultured cells, and tissue using the Applied Biosystems™ MagMAX™ High Molecular Weight (HMW) DNA Kit on the Thermo Scientific™ KingFisher™ Flex system. The fully automated workflow-comprising cell lysis, magnetic-bead-based DNA binding, sequential washing, and low-salt elution-reduces manual intervention and potential shearing events while improving consistency across users and sample types. An optional brief enzymatic digestion (<15 min) was included for tissue samples to optimize recovery of long fragments. DNA yield and integrity were quantified using Qubit™ fluorometry, Agilent TapeStation, and pulsed-field electrophoresis, and performance was compared against commonly used silica column- and precipitation-based kits. Long-read sequencing was used to assess read length distribution, coverage uniformity, and structural variant detection. Legacy manual and semi-automated gDNA kits exhibited substantial variability in yield (coefficient of variation (CV) >15%) and produced fragmented DNA with median fragment sizes typically below 20 kb, resulting in truncated reads and reduced sensitivity for detecting large deletions and rearrangements. In contrast, the MagMAX™ HMW automated workflow consistently produced DNA fragments >100 kb, with an average yield of 5 µg per sample and CV <1 across replicates; across sample types, 85% of fragments exceeded 40 kb and 75% exceeded 100 kb, enabling longer read lengths and higher variant concordance. The automated process supported batch processing of up to 96 samples in under two hours with less than 30 minutes of hands-on time, substantially reducing operator bias and inter-run variability. Overall, automating the MagMAX™ HMW DNA kit on the KingFisher™ instrument proved essential for preserving DNA integrity, improving reproducibility, and increasing throughput, enabling generation of high-quality HMW DNA that enhances read continuity, structural variant resolution, and data reliability-supporting more complete genomic profiling and accelerating translational oncology research.