Efficient small DNA fragment sequencing and miRNA, small RNA or csRNA-seq libraries using AVITI

Next-Generation Sequencing (NGS) catalyzed breakthroughs across various scientific domains. Illumina's sequencing by synthesis method has long been essential for NGS but emerging technologies like Element Biosciences' sequencing by avidity (AVITI) represent a novel approach. It has been reported that AVITI offers improved signal-to-noise ratios and cost reductions. However, the method relies on rolling circle amplification which can be impacted by polymer size, raising questions about its efficacy sequencing small RNAs (sRNAs) libraries including microRNAs (miRNAs), piwi-interacting RNAs (piRNAs), and others that are crucial regulators of gene expression and involved in various biological processes. In addition, capturing capped small RNAs (csRNA-seq) has emerged as a powerful method to map active or “nascent” RNA polymerase II transcription initiation in tissues and clinical samples. Finally, we present an accelerated and optimized protocol for small fragment sequencing with AVITI that aligns seamlessly with available sRNA library kits to meet this need. Capped small (cs)RNA-sequencing for human A375 and BT474 cells, as well as blood samples from cattle and bison using Illumina and AVITI sequencing.