A zinc finger protein array for the visual detection of specific DNA sequences for diagnostic applications

Nucleic Acids Res. 2011 Mar;39(5):e29. doi: 10.1093/nar/gkq1214. Epub 2010 Dec 5.

Abstract

The visual detection of specific double-stranded DNA sequences possesses great potential for the development of diagnostics. Zinc finger domains provide a powerful scaffold for creating custom DNA-binding proteins that recognize specific DNA sequences. We previously demonstrated sequence-enabled reassembly of TEM-1 β-lactamase (SEER-LAC), a system consisting of two inactive fragments of β-lactamase each linked to engineered zinc finger proteins (ZFPs). Here the SEER-LAC system was applied to develop ZFP arrays that function as simple devices to identify bacterial double-stranded DNA sequences. The ZFP arrays provided a quantitative assay with a detection limit of 50 fmol of target DNA. The method could distinguish target DNA from non-target DNA within 5 min. The ZFP arrays provided sufficient sensitivity and high specificity to recognize specific DNA sequences. These results suggest that ZFP arrays have the potential to be developed into a simple and rapid point-of-care (POC) diagnostic for the multiplexed detection of pathogens.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • DNA, Bacterial / chemistry
  • DNA-Binding Proteins / analysis
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Escherichia coli / genetics
  • Molecular Diagnostic Techniques
  • Protein Array Analysis / methods*
  • Protein Engineering
  • Sequence Analysis, DNA*
  • Zinc Fingers*
  • beta-Lactamases / genetics

Substances

  • DNA, Bacterial
  • DNA-Binding Proteins
  • beta-Lactamases
  • beta-lactamase TEM-1