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The importance of different sources and sequencing protocols in increasing the accuracy of NGS analysis in diagnostic applications
Kvapilová, Kateřina ; Kozmik, Zbyněk (advisor) ; Piherová, Lenka (referee) ; Slabý, Ondřej (referee)
1 ABSTRACT Continued advances in next-generation sequencing (NGS) technology, such as capacity, speed, and reduced cost per sequenced base, revolutionize personalized medicine and bring genomics into routine clinical practice. Nevertheless, NGS is still under rapid development, and the variability of sequencing protocols and validation procedures is one of its current bottlenecks. This thesis aimed to study the influence of different sample sources and NGS protocols (NGS library construction-sequencing-data analysis) on the accuracy of NGS analysis in diagnostic applications. In the first study, performed during the COVID-19 pandemic outbreak, we developed NGS protocols suitable for a whole genome analysis of the SARS-CoV-2 virus. Subsequently, in the second study, we examined the suitability of human saliva-derived gDNA for genomic/genetic analysis of selected variant types compared to traditional blood-derived gDNAusing validated NGS protocol and statistical comparison of the obtained data. Whole genome analysis of the SARS-CoV-2 genome was performed using two captured- based approaches and one amplicon-based approach to study the quality and effectivity of the respective NGS protocols. Synthetic controls were employed to verify the accuracy and specificity of the developed NGS protocols. We proved that...
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