SplintQuant: A method for accurately quantifying circular RNA transcript abundance without reverse transcription bias

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Reverse transcription of RNA is fallible, introducing biases and confounding the quantification of transcript abundance. We demonstrate that circular RNAs (circRNAs) are more subjective to overestimation of transcript abundance than cognate linear RNAs due to their covalently closed, circular form, producing multiple concatameric products from a single priming of reverse transcriptase. We developed SplintQuant, where custom DNA oligonucleotides are ligated by PBCV-1 DNA ligase only when bound to their target RNA. These circRNA-specific DNA oligonucleotides are terminally tagged with universal primers, allowing SplintQuant to accurately quantify even lowly abundant circRNAs through highly specific quantitative PCR (qPCR) in the absence of reverse transcription. SplintQuant is sensitive, specific, highly reproducible, and applicable to the quantification of canonical and noncanonical RNA transcripts including alternative splice variants, gene fusions, and offers a gold-standard approach for accurately quantifying circRNAs.

Original languageEnglish
Pages (from-to)1202-1210
Number of pages9
JournalRNA
Volume25
Issue number9
DOIs
Publication statusPublished - 1 Jan 2019

Keywords

  • Circular RNA
  • QPCR
  • Reverse transcription
  • Splint ligation

Fingerprint Dive into the research topics of 'SplintQuant: A method for accurately quantifying circular RNA transcript abundance without reverse transcription bias'. Together they form a unique fingerprint.

  • Cite this