Accurate Quantity and Quality Analysis of NGS Library Preparations using the Fragment Analyzer™

Labs doing NGS library prep are confronted with many challenges each day including varying incoming sample quality, sample prep loss, pooling/barcoding issues, quantification inconsistencies and fluctuating sample numbers to... [ view full abstract ]

Labs doing NGS library prep are confronted with many challenges each day including varying incoming sample quality, sample prep loss, pooling/barcoding issues, quantification inconsistencies and fluctuating sample numbers to name a few.   Propitiously, the major sequencing platforms have enhanced their capabilities allowing the use of suboptimal, small sized and/or low quantity DNA and RNA raw materials.  This in turn has opened up new and previously unimagined ways to generate sequencing data.  With these advances, however, inaccurate quantification of library preps made from poor quality raw materials, from pooled libraries made from different samples or unbalanced barcoding strategies can affect both sequencing results and radically affect costs.  Back in the early days of NGS, few NGS library prep methods were available. Today there are many ways to produce libraries.  A better understanding of the raw and finished nucleic acid materials are needed because labs face both constraining challenges imposed on them by working with low quality nucleic acids and enabling challenges that open the possibility of doing longer fragment length reads.    Due to these factors, instrumentation which can sensitively and accurately assess the quality and quantity of nucleic acids can improve the consistency of results generation.  The Fragment Analyzer™ Automated CE System from Advanced Analytical was purposely designed to meet the challenges of NGS library prep assessment.  By utilizing the superior properties of capillary electrophoresis, the Fragment Analyzer can effectively separate fragment sizes from very small (microRNA) to very large (genomic DNA) and all sizes in between as well as provide high quantification accuracy.  Information will be presented on why accurate concentration and size assessment is important to achieving good sequencing data and final results derived from both short and long read sequencing platforms.