Arrest of rolling circle amplification by protein-binding DNA aptamers

Chemistry. 2014 Feb 24;20(9):2420-4. doi: 10.1002/chem.201304292. Epub 2014 Feb 7.

Abstract

Certain DNA polymerases, such as ϕ29 DNA polymerase, can isothermally copy the sequence of a circular template round by round in a process known as rolling circle amplification (RCA), which results in super-long single-stranded (ss) DNA molecules made of tandem repeats. The power of RCA reflects the high processivity and the strand-displacement ability of these polymerases. In this work, the ability of ϕ29DNAP to carry out RCA over circular templates containing a protein-binding DNA aptamer sequence was investigated. It was found that protein-aptamer interactions can prevent this DNA polymerase from reading through the aptameric domain. This finding indicates that protein-binding DNA aptamers can form highly stable complexes with their targets in solution. This novel observation was exploited by translating RCA arrest into a simple and convenient colorimetric assay for the detection of specific protein targets, which continues to showcase the versatility of aptamers as molecular recognition elements for biosensing applications.

Keywords: aptamers; biosensors; molecular competition; protein detection; rolling circle amplification.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aptamers, Nucleotide / biosynthesis*
  • Aptamers, Nucleotide / chemistry*
  • DNA-Binding Proteins / chemistry*
  • DNA-Directed DNA Polymerase / chemistry*
  • Nucleic Acid Amplification Techniques / methods
  • Oligonucleotides / biosynthesis*
  • Oligonucleotides / chemistry*

Substances

  • Aptamers, Nucleotide
  • DNA-Binding Proteins
  • Oligonucleotides
  • DNA-Directed DNA Polymerase