Characterization and mitigation of fragmentation enzyme-induced dual stranded artifacts

Area of Science:

• Genomics
• Molecular Biology
• Bioinformatics

Background:

• High-throughput sequencing requires DNA fragmentation.
• Enzyme cocktails offer a cost-effective alternative to acoustic shearing for DNA fragmentation.
• Enzyme-based fragmentation can introduce artifacts into sequencing libraries.

Purpose of the Study:

• To identify and characterize artifacts generated by enzyme-based DNA fragmentation.
• To develop a computational tool for detecting and removing these artifacts.
• To assess the impact of these artifacts on downstream genomic analyses.

Main Methods:

• Development of Fragmentation Artifact Detection and Elimination (FADE) software.
• Analysis of sequencing data to identify artifact-derived reads.
• Evaluation of FADE's performance in removing artifacts.
• Assessment of artifact impact on targeted resequencing and rare variant discovery.

Main Results:

• Enzyme-based fragmentation creates library molecules with regions from opposite DNA strands.
• These artifacts can lead to false variant calls at low allele frequencies (<5%).
• FADE effectively removes artifact-derived reads and mitigates downstream analysis biases.

Conclusions:

• Enzyme-based DNA fragmentation methods introduce specific artifacts.
• FADE is a crucial tool for accurate variant calling, especially in sensitive applications.
• Mitigating these artifacts is essential for reliable genomic data interpretation.