期刊
NUCLEIC ACIDS RESEARCH
卷 43, 期 12, 页码 6125-6133出版社
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkv519
关键词
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资金
- Canadian Institutes of Health Research (CIHR)
- Ontario Ministry of Research Innovation
- Natural Sciences and Engineering Research Council (NSERC) of Canada (Discovery Grant and Strategic Project Grant)
- CIHR
- NSERC of Canada
In vitro selection of RNA-cleaving DNAzymes is a powerful method for isolating metal-specific DNA. A few successful examples are known, but it is still difficult to target some thiophilic metals such as Cd2+ due to limited functional groups in DNA. While using modified bases expands the chemical functionality of DNA, a single phosphorothioate modification might boost its affinity for thiophilic metals without complicating the selection process or using bases that are not commercially available. In this work, the first such in vitro selection for Cd2+ is reported. After using a blocking DNA and negative selections to rationally direct the library outcome, a highly specific DNAzyme with only 12 nucleotides in the catalytic loop is isolated. This DNAzyme has a cleavage rate of 0.12 min(-1) with 10 mu M Cd2+ at pH 6.0. The R-p form of the substrate is cleaved similar to 100-fold faster than the S-p form. The DNAzyme is most active with Cd2+ and its selectivity against Zn2+ is over 100 000-fold. Its application in detecting Cd2+ is also demonstrated. The idea of introducing single modifications in the fixed region expands the scope of DNA/metal interactions with minimal perturbation of DNA structure and property.
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