Exploring the impact of primer length on efficient gene detection via high-throughput sequencing

• DOI: 10.1038/s41467-024-49685-0
• Publication Date: 2024-07-12T11:02:16Z
• Authors: Julia Micheel; Aram Safrastyan; Franziska Aron; Damian Wollny
• Keywords: Primer (cosmetics)

Abstract Reverse transcription (RT) is a crucial step in most RNA analysis methods. Optimizing protocols for this initial stage critical effective target detection, particularly when working with limited input RNA. Several factors, such as the material quality and reaction conditions, influence RT efficiency. However, effect of primer length on gene detection efficiency remains largely unknown. Thus, we investigate its impact by generating RNA-seq libraries random primers 6, 12, 18, or 24 nucleotides. To our surprise, 18mer shows superior overall transcript compared to commonly used 6mer primer, especially detecting longer transcripts complex human tissue samples. This study highlights role efficiency, which has significant potential benefit various transcriptomic assays, from basic research clinical diagnostics, given central RNA-related analyses.