Ultrasensitive detection of miRNA via one-step rolling circle-quantitative PCR (RC-qPCR)

Mingcheng Xu, Jiawei Ye, Dan Yang, Abdu Ahmed Abdullah AL-maskri, Haihong Hu, Cheulhee Jung, Sheng Cai, Su Zeng

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

A novel microRNA (miRNA) quantification method has been developed using one-step rolling circle-quantitative PCR (RC-qPCR) analysis. Vent (exo-) DNA polymerase is firstly utilized to combine a rolling circle amplification (RCA) and qPCR in one step with high sensitivity and specificity in our RC-qPCR assay. Before performing the RC-qPCR, a padlock probe is ligated only when it is perfectly hybridized with miRNA. This ligation-based miRNA assay is highly specific for mature miRNAs, discriminating among related miRNAs that differ by as little as one nucleotide. It exhibits a dynamic range of seven orders of magnitude with a detection limit of 500 aM, and could be also used for the quantification of other small RNA molecules such as short interfering RNAs (siRNAs).

Original languageEnglish
JournalAnalytica Chimica Acta
DOIs
Publication statusPublished - 2019 Jan 1

Keywords

  • MiR-200a
  • MiRNA detection
  • One-step amplification
  • Rolling circle amplification
  • Rolling circle-quantitative PCR (RC-qPCR)
  • Vent (exo-) DNA polymerase

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Environmental Chemistry
  • Spectroscopy

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