Invader probes: harnessing the energy of intercalation to facilitate recognition of chromosomal DNA for diagnostic applications
|
| |
Authors: | Dale C Guenther Grace H Anderson Saswata Karmakar Brooke A Anderson Bradley A Didion Wei Guo John P Verstegen Patrick J Hrdlicka |
| |
Institution: | a Department of Chemistry , University of Idaho , 875 Perimeter Dr , Moscow , ID 83844-2343 , USA . Email: ; b Department of Biological Sciences , Montana Tech of the University of Montana , 1300 W Park St , Butte , MT 59701-8997 , USA ; c MoFA , PO Box 930187, 419 Venture Ct. , Verona , WI 53593 , USA |
| |
Abstract: | Development of probes capable of recognizing specific regions of chromosomal DNA has been a long-standing goal for chemical biologists. Current strategies such as PNA, triplex-forming oligonucleotides, and polyamides are subject to target choice limitations and/or necessitate non-physiological conditions, leaving a need for alternative approaches. Toward this end, we have recently introduced double-stranded oligonucleotide probes that are energetically activated for DNA recognition through modification with +1 interstrand zippers of intercalator-functionalized nucleotide monomers. Herein, probes with different chemistries and architectures – varying in the position, number, and distance between the intercalator zippers – are studied with respect to hybridization energetics and DNA-targeting properties. Experiments with model DNA targets demonstrate that optimized probes enable efficient (C
50 < 1 μM), fast (t
50 < 3 h), kinetically stable (>24 h), and single nucleotide specific recognition of DNA targets at physiologically relevant ionic strengths. Optimized probes were used in non-denaturing fluorescence in situ hybridization experiments for detection of gender-specific mixed-sequence chromosomal DNA target regions. These probes present themselves as a promising strategy for recognition of chromosomal DNA, which will enable development of new tools for applications in molecular biology, genomic engineering and nanotechnology. |
| |
Keywords: | |
|
|