Electronic detection of DNA mutation based on strand exchange reaction.

A new approach to electronic detection of a single base mismatch is described. The assay involves the electrochemical measurements of DNA strand exchange reactions (SERs) between electrode-bound redox-modified DNA duplex and target DNA, where the sequence of redox-modified DNA is exchangeable to that of the target DNA. The presence of a single base mismatch can be determined from the slower SER rates compared with fully matched DNA.

Development of pyrrole-imidazole polyamide for specific regulation of human aurora kinase-A and -B gene expression.

Pyrrole-imidazole polyamide (PIP) is a nuclease-resistant novel compound that inhibits gene expression through binding to the minor groove of DNA. Human aurora kinase-A (AURKA) and -B (AURKB) are important regulators in mitosis during the cell cycle. In this study, two specific PIPs (PIP-A and PIP-B) targeting AURKA and AURKB promoter regions were designed and synthesized, and their biological effects were investigated by several in vitro assays. PIP-A and PIP-B significantly inhibited the promoter activities, mRNA expression, and protein levels of AURKA and AURKB, respectively, in a concentration-dependent manner. Moreover, 1:1 combination treatment with both PIPs demonstrated prominent antiproliferative synergy (CI value [ED(50)] = 0.256) to HeLa cells as a result of inducing apoptosis-mediated severe catastrophe of cell-cycle progression. The novel synthesized PIP-A and PIP-B are potent and specific gene-silencing agents for AURKA and AURKB.

Target-induced strand release (TISR) from aptamer-DNA duplex: a general strategy for electronic detection of biomolecules ranging from a small molecule to a large protein.

We describe a general strategy for electronic aptamer-based biosensor that is based on target-induced strand release (TISR) of a redox-modified aptamer from the aptamer-capture DNA duplex bound on an electrode.

2'-anthraquinone-conjugated oligonucleotide as an electrochemical probe for DNA mismatch.

We previously prepared the oligonucleotides (ODNs) conjugated to an anthraquinone (AQ) group via one carbon linker at the 2'-sugar position. When these modified ODNs bind to cDNA sequences, the AQ moiety can be intercalated into the predetermined base-pair pocket of a duplex DNA. In this paper, 2'-AQ-modified ODNs are shown to be an excellent electrochemical probe to clarify the effect of a mismatch base on the charge transfer (CT) though DNA. Two types of DNA-modified electrodes were constructed by assembly of disulfide-terminated 2'-AQ-ODN duplexes onto gold electrodes. One type of electrodes (system I) contains fully matched base pairs or a single-base mismatch in duplex DNA between the redox center and the electrode. The other (system II) consists of the mismatch but at the outside of the redox center. The modified electrodes were analyzed by cyclic voltammetry to estimate the CT rate through duplex DNA. In system I, the CT rate was found to be approximately 50 s (-1) for the fully matched AQ-ODN duplexes, while the CT rates of the mismatched DNA were considerably slower than that of the fully matched DNA. In system II, the AQ-ODN duplexes showed almost similar CT rates ( approximately 50 s (-1)) for the fully matched DNA and for the mismatched DNAs. The detection of a single-base mismatch was then performed by chronocoulometry (CC). All the DNA duplexes containing a mismatch base in system I gave the reduced electrochemical responses when compared to the fully matched DNA. In particular, the mismatched DNAs including G--A mismatch can be differentiated from fully matched DNA without using any electrochemical catalyst. We further tested the usefulness of single-stranded (ss) AQ-ODN immobilized on a gold electrode in the electrochemical detection of a single-base mismatch through hybridization assay. The ss-AQ-ODN electrodes were immersed in target-containing buffer at room temperature, and the CC measurements were carried out to see the changes in the integrated charge. Within 60 min, the mismatched DNA was clearly distinguishable by the CC differences from the fully matched target. Thus, the electrochemical hybridization assay provides an easy and convenient detection for DNA mutation that does not require any extra reagents, catalyst, target labeling, and washing steps.

Electrochemical properties of anthraquinone-capped DNA-hairpins immobilized on gold surface.

Disulfide functionalized hairpin-forming DNAs having an anthraquinone linker as a redox probe with variable number of A-T base pairs were synthesized. The electron transfer through the DNAs in self-assembled monolayers on gold surface was studied.

Electrochemical biosensors based on DNA strand exchange.

Electrochemical biosensors for the detection of specific DNA sequences have been described. The redox-modified DNA sequences were hybridized with the 3'-thiol-modified DNA sequences as capture DNA strand. The resulting duplexes were assembled on a gold electrode surface to make DNA chip. Upon addition of fully complementary DNA or mismatch-containing DNA, the time-dependent changes in redox signal were monitored. Mismatched DNA gave less than 20% change when compared with fully matched DNA under the same conditions. The present assay format is thus applicable in discrimination of mutated DNA from fully matched DNA.

13C NMR longitudinal relaxation time studies of a molecular tweezers derived from a calixarene-porphyrin conjugate.

Zn(II)porphyrin-substituted calix[4]arene 1 serves as molecular tweezers for 1,4-diazabicyclo[2.2.2] octane (DABCO) selectively, which led to the formation of Ensemble I. The molecular segments composing the calixarene cavity change upon inclusion of DABCO as Ensemble I were evaluated through (13)C NMR longitudinal relaxation times (T(1)) for the first time. As for Ensemble I, the 1:1 complex should be formed. The T(1) values for Ensemble I are generally smaller than those for 1: in CDCl(2)CDCl(2), DT1 = 5.03 s for C-1, 5.31 s for C-2, 0.13 s for C-3, 0.7 s for C-4, and 0.16 s for C-5. This substantiates that the rings of Ensemble I are firmly freezed because of the two-point coordination by DABCO. In 1, the T1 values for C-3 are always greater than those for C-4, and the difference between C-3 and C-4 is slight. As for Ensemble I, on the other hand, the difference between C-3 and C-4 is large. We can suggest two different motions for phenol units in 1 and Ensemble I: a rotational motion around a C-1 to C-4 axis (A) and a seesaw motion around a C-2 to C-2' axis (B). The data indicate that in Ensemble I motion (A) is predominant over motion (B). This indicates that motion (B) is specifically suppressed because of the two-point coordination interactions in Ensemble I.

Electrochemical detection of DNA single base mismatch by the use of strand exchange reaction.

The electrochemical method based on the strand exchange reaction (SER) for the detection of DNA single-base mismatch has been developed. Different electrochemical responses due to the slower SER rates for mismatch containing DNA than fully matched DNA were observed by using the redox-modified partial duplex DNA immobilized on gold electrode as an analytical probe.

Electrochemical detection of single base mismatches in DNA using redox-modified oligonucleotides.

A facile synthesis of new anthraquinone (AQ)-modified oligonucleotides was described. The AQ-oligomer-bound electrode was used for the electrochemical detection of a single base mismatch in DNA.

Probing DNA sequences by anthraquinone-modified DNA that is immobilized on gold surface.

2'-Anthraquinone-modified oligonucleotide (AQ-ODN) possessing disulfide terminus has been immobilized on the gold electrode surface. The AQ-ODN-modified electrode showed faster electron transferability in double-stranded form than that in single-stranded form.