Application of the Agilent 2100 Bioanalyzer instrument as quality control for next-generation sequencing

Area of Science:

• Forensic Science
• Molecular Biology
• Genetics

Background:

• Next-generation sequencing (NGS) advances forensic DNA analysis but faces challenges in complex sample processing and library preparation.
• Accurate quantification and quality assessment of DNA libraries are critical for optimal sequencing performance, especially with degraded samples.

Purpose of the Study:

• To evaluate the Agilent 2100 Bioanalyzer's utility for quality control in forensic DNA library preparation for NGS.
• To assess the performance of the Agilent 2100 Bioanalyzer with challenging, degraded postmortem samples.

Main Methods:

• Analyzed DNA concentrations, fluorescence unit (FU) values, and adapter dimer concentrations in libraries from buccal swabs and decomposed human tissue.
• Conducted a serial dilution sensitivity study using buccal swab DNA to assess the impact of input concentration on measurements.
• Examined repeatability and quantification accuracy of the Agilent 2100 Bioanalyzer across multiple runs.

Main Results:

• Sensitivity analysis showed decreased FU values and increased adapter dimers with lower DNA input.
• Observed variability in total DNA concentrations and peak heights indicated repeatability challenges in quantification.
• Successfully detected adapter dimers in degraded samples, highlighting their importance for NGS library quality.

Conclusions:

• The Agilent 2100 Bioanalyzer is a valuable tool for assessing DNA library quality in forensic NGS applications.
• Detecting adapter dimers is crucial for ensuring successful NGS library preparation, particularly with degraded forensic samples.
• While quantification repeatability needs consideration, the Bioanalyzer aids in critical quality control aspects for forensic DNA analysis.