Preparation of Carbodiimide-terminated Dithiolane Self-Assembly Monolayers as a New DNA-Immobilization Method

Abstract

Gene-detecting methods using a DNA-fixed substrate are important in biotechnology and related fields. DNA microarray or DNA chip based on a DNA-immobilized substrate have been developed to enable a high-throughput analysis of many genes simultaneously. [1][2][3][4][5][6][7][8][9] Many kinds of DNA fragments, called a capture DNA probe, are immobilized on the substrate of a DNA microarray or DNA chip. A proper DNA fragment in a sample can hybridize with the complementary capture DNA probe on the substrate, and its hybrid can be detected by the combination of DNA detecting reagents. Such a system has been applied to a study of gene expression and mutation analysis including single nucleotide polymorphism (SNP) analysis.1-3,6-9 A goldcovered substrate, having conductivity sufficient for electrochemical DNA detection, is also a good candidate for a DNA chip of the next generation from the viewpoint of developing a simple, quick, and miniaturized detection system and in fact, many papers have dealt with this subject. [10][11][12][13][14][15] To construct such a system, it is important to develop a reproducible method to immobilize a capture DNA probe on the gold surface, and many kinds of immobilization methods have been reported. [16][17][18][19][20][21][22] An immobilization method through the Au-S linkage is one of the conventional methods. It is known that thiolated DNA can bind to a gold surface, and detailed studies, such as an estimation of the density of DNA immobilization, were carried out. [16][17][18][19] In addition to the direct immobilization of thiolated DNA on a gold surface, many kinds of indirect immobilization methods were reported.Self-assembly monolayers (SAMs) were prepared by an alkanethiol carrying a functional group at the opposite terminus, and DNA was covalently attached through it; for example, DNA carrying an amino terminus can be attached to carboxylic acid-terminated SAMs using a condensation reagent or active ester derivatives. 20,21 Recently, Childsey et al. reported a novel immobilization method for acetylated DNA on a SAM surface having an azide group. 22 Such a SAM system can avoid nonspecific absorption by introducing an additional hydroxyl alkanethiol, such as 6-mercaptohexanol (6MH). An immobilization method based on biotin-avidin interaction, where biotinyl DNA was immobilized on an avidin-covered gold electrode, was also reported. 23,24 Although many of these DNA-immobilization methods are based on a SAM system, all of them require some sort of chemical modification of DNA, resulting in a time-consuming procedure with a low modification yield of DNA.In this paper, we developed a simple DNA-immobilization method for intact DNA, as shown in Fig. 1. We designed and synthesized compound 1 carrying a dithiolane and carbodiimide part at its termini. Ditholane derivatives are known to be immobilized on a gold surface through the Au-S linkage, 25,26 and have been applied to immobilize DNA or protein on a gold surface. 27 Carbodiimide derivatives are known to react with the imino moie...

Biochemical Assays of Immobilized Oligonucleotides with Mass Spectrometry

“…This report provides a straightforward strategy for implementing a broad class of assays that use immobilized oligonucleotides. The common use of self-assembled monolayer of alkanethiolates on gold as substrates for the preparation of oligonucleotide arrays – and the availability of commercial instruments for MALDI-TOF MS that can acquire of order 1000 spectra/h provide a direct route to pursuing applications of arrays of oligonucleotides. The examples in this paper extend the SAMDI MS method to biochemical assays of oligonucleotides, increasing the scope of this method to a broad range of applications in interfacial chemistry and biology …”

Synthesis, Characterization, and Antiproliferative Activities of Novel Ferrocenophanic Suberamides against Human Triple-Negative MDA-MB-231 and Hormone-Dependent MCF-7 Breast Cancer Cells

“…The first examples of ferrocenic compounds exerting antiproliferative activity against cancer cells were ferricenium derivatives, albeit with low activity . Since then, ferrocene has been considered for the development of compounds with biological applications such as anticancer agents, antimalarial agents, DNA detection, and enzymatic inhibition against HIV (such as topoisomerase and integrase inactivation).…”

Fabrication of patterned surfaces by photolithographic exposure of DNA hairpins carrying a novel photolabile group

“…Hybrid top-down/bottom-up processes, which combine the precision of lithographic techniques and the parallelism of self-assembly have been actively pursued in order to create patterns on silicon surfaces functionalised with self-assembled monolayers (SAMs) [5][6][7]. However, the ability to organise biomolecules, such as DNA on surfaces, such as glass [8,9] and gold [10][11][12] is an area of increased interest.…”

“…One method to create these patterns is by exposure of thiol-stabilised gold NPs supported on silicon wafers to UV light leading to oxidation of the thiol and coagulation of the NPs [20], or photo-crosslinking of C60 derivatives [21], forming dense structures that are resistant to removal by organic solvents. Methods exist to further derivatise gold surfaces, such as those formed, with oligonucleotide sequences [11,22,23]. Alternatively it is possible to modify a specific region of a surface introducing chemical functionality to direct the adsorption of particulate species.…”