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Sequence‐Specific Covalent Capture Coupled with High‐Contrast Nanopore Detection of a Disease‐Derived Nucleic Acid Sequence
Author(s) -
Nejad Maryam Imani,
Shi Ruicheng,
Zhang Xinyue,
Gu LiQun,
Gates Kent S.
Publication year - 2017
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201700204
Subject(s) - nucleic acid , molecular beacon , sequence (biology) , nanopore , dna , chemistry , covalent bond , oligonucleotide , biochemistry , biology , microbiology and biotechnology , combinatorial chemistry , nanotechnology , materials science , organic chemistry
Hybridization‐based methods for the detection of nucleic acid sequences are important in research and medicine. Short probes provide sequence specificity, but do not always provide a durable signal. Sequence‐specific covalent crosslink formation can anchor probes to target DNA and might also provide an additional layer of target selectivity. Here, we developed a new crosslinking reaction for the covalent capture of specific nucleic acid sequences. This process involved reaction of an abasic (Ap) site in a probe strand with an adenine residue in the target strand and was used for the detection of a disease‐relevant T→A mutation at position 1799 of the human BRAF kinase gene sequence. Ap‐containing probes were easily prepared and displayed excellent specificity for the mutant sequence under isothermal assay conditions. It was further shown that nanopore technology provides a high contrast—in essence, digital—signal that enables sensitive, single‐molecule sensing of the cross‐linked duplexes.