Improved characterization of 3' single-cell RNA-seq libraries with paired-end avidity sequencing.

ABSTRACT: Prevailing poly(dT)-primed 3' single-cell RNA-seq protocols generate barcoded cDNA fragments containing the reverse transcriptase priming site or in principle the polyadenylation site. Direct sequencing across this site was historically difficult because of DNA sequencing errors induced by the homopolymeric primer at the 'barcode' end. Here, we evaluate the capability of 'avidity base chemistry' DNA sequencing from Element Biosciences to sequence through the primer and enable accurate paired-end read alignment and precise quantification of polyadenylation sites. We find that the Element Aviti instrument sequences through the thymine homopolymer into the subsequent cDNA sequence without detectable loss of accuracy. The additional sequence enables direct and independent assignment of reads t

SUBMITTER: Chamberlin JT 

PROVIDER: S-EPMC11655283 | biostudies-literature | 2024 Dec

REPOSITORIES: biostudies-literature