Precision thermal cyclers ensure accurate NGS DNA library preparation.
Sponsored Content by Analytik Jena USReviewed by Andrea SalazarOct 16 2024 Next-generation sequencing is a massively parallel sequencing technology that provides various applications, including thorough analysis of entire genomes or select target sections.1
In all NGS applications, the quality of DNA sequencing libraries significantly impacts the preparation processes and, consequently, the result.2 It is, therefore, critical to undertake DNA library preparation with maximum precision and reproducibility. This article presents the preparation of DNA amplicon sequencing libraries for NGS using the NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® on the Biometra TAdvanced 96 SG. It also describes library quantification using the real-time thermal cycler qTOWERiris .
Table 1. Four gene targets for amplicon library preparation were chosen derived from human genomic DNA representing varying amplicon lengths and GC content. Source: Analytik Jena USInstrumentation Library preparation workflow The NEB Next Ultra™ II DNA Library Prep Kit for Illumina® was used to prepare a library from purified PCR products ranging from 300 ng to 1 µg. All procedures were followed from the handbook.
Library quantification The resulting libraries were quantified using the real-time thermal cycler qTOWERiris using the sample configuration and PCR software , as specified in the NEBNext® Library Quant Kit for Illumina® manual. Results and discussion The sample libraries were processed consistently across all library preparation processes, regardless of their position on the Biometra TAdvanced 96 SG’s heat block, as seen by gel electrophoresis following adaptor ligation and PCR enrichment using Index Primers Set1 .
In conclusion, the workflow presented, utilizing PCR and real-time PCR thermal cyclers from Analytik Jena and a library preparation kit from NEB, proved to be a reliable and efficient method for NGS DNA library preparation for amplicon sequencing, regardless of gene target, amplicon length, or GC content.
The Biometra TAdvanced thermal cycler works well with the NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® to produce consistent DNA amplicon libraries. The resultant libraries had variable lengths and GC content, demonstrating the equipment and reagents’ appropriateness for this procedure. About Life Science The Life Science product area demonstrates the biotechnological competence of Analytik Jena AG. We provide a wide product spectrum for automated total, as well as individual solutions for molecular diagnostics. Our products are focused to offer you a quality and the reproducibility of your laboratory results. This will surely ease your daily work and speed up your work processes in a certain way.
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