Immobilization of DNA via oligonucleotides containing an aldehyde or carboxylic acid group at the 5' terminus

Kremsky, Jonathan N.; Wooters, Joseph; Dougherty, Joseph P.; Meyers, Rachel; Collins, Mary; Brown, Eugene L.

doi:10.1093/nar/15.7.2891

Cited by 103 publications

(80 citation statements)

References 33 publications

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“…At a high voltage (30 V/cm), the ARFI of spots whose locations were far from the cathode was stronger than those near the cathode. The ARFI of spots furthest from the cathode was 2.05610 4 , which was 4-fold higher than that of the tenth column spots located close to the cathode, which was 5.06610 3 . At a lower voltage (5 V/cm), the ARFI of the first column spots was 3.89610 4 , and this value was 1.14-fold higher than the nearest one (the tenth column spots, 3.41610 4 ).…”

Section: Figurementioning

confidence: 72%

“…2B. The ARFI of the control spots was only 981 even when the readings were scanned under a scanner setting of 90% laser power and 90% photomultiplier tube (PMT) gain; the corresponding value for the oligonucleotides was 2.48610 4 . Although the fluorescence intensity of oligonucleotides was decreased, the S/N increased from 1.14 to 25.2.…”

Section: Electrophoresis For Reducing the Nonspecifically Bound Targementioning

confidence: 99%

“…6. When electrophoresis was carried out at 5 V/cm and at room temperature (227C) for 30 min, the ARFIs of P 0 , P 1 , P 2 , P 3 and P were 3.54610 4 , 1.84610 4 , 879, 318 and 294, respectively (Figs. 6A, C).…”

Section: Hybridization Of Immobilized Oligonucleotidesmentioning

confidence: 99%

“…The yield of nucleic acids immobilization and the efficiency of hybridization are key factors to the accuracy and sensitivity of the microarrays prepared by spotting. A number of chemical immobilization approaches have been developed to attach nucleic acids on various substrates, such as glass [4][5][6], silicon [7], and plastics [8,9]. Because of the low immobilization efficiency of nucleic acids in most of these methods, purification of nucleic acids (either synthesized probes or PCR products) by commercial purification cartridges is required to achieve high quantity in microarray spotting [10].…”

Section: Introductionmentioning

confidence: 99%

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An improved gel‐based DNA microarray method for detecting single nucleotide mismatch

et al. 2006

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3-D polyacrylamide gel-based DNA microarray platforms provide a high capacity for nucleic acids immobilization and a solution-mimicking environment for hybridization. However, several technological bottlenecks still remain in these platforms, such as difficult microarray preparation and high fluorescent background, which limit their application. In this study, two new approaches have been developed to improve the convenience in microarray preparation and to reduce the background after hybridization. To control the polymerization process, solutions containing acrylamide-modified oligonucleotide, acrylamide, glycerol and ammonium persulfate are spotted onto a functionalized glass slide, and then the slide is transferred to a vacuum chamber with TEMED, so that TEMED is vaporized and diffused into the spots to induce polymerization. By applying an electric field across a hybridized microarray to remove the nonspecifically bound labeled targets, this approach can solve the problem of high fluorescent background of the gel-based microarray after hybridization. Experimental results show that our immobilization method can be used to construct high quality microarrays and exhibits good reproducibility. Moreover, the polymerization is not affected by PCR medium, so that PCR products can be used for microarray construction without being treated by commercial purification cartridges. Electrophoresis can improve the signal-to-noise significantly and has the ability to differentiate single nucleotide variation between two homozygotes and a heterozygote. Our results demonstrated that this is a reliable novel method for high-throughput mutation analysis and disease diagnosis.

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Section: Figurementioning

confidence: 72%

Section: Electrophoresis For Reducing the Nonspecifically Bound Targementioning

confidence: 99%

Section: Hybridization Of Immobilized Oligonucleotidesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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An improved gel‐based DNA microarray method for detecting single nucleotide mismatch

et al. 2006

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“…The principal advantages of using surfacebound oligonucleotides over those in solution include ease of purification, conservation of material and reagents, reduction of interference between oligonucleotides and facilitated sample handling (12). Previously explored surfaces for immobilization of DNA include latex beads (5), polystyrene (1), carbon electrodes (22)(23)(24)(25), gold (17,23,(26)(27)(28) and oxidized silicon or glass (3,8,11,(29)(30)(31)(32). The surface chemistries involved with these substrates do not generally possess all of the desired characteristics of an ideal surface.…”

Section: Introductionmentioning

confidence: 99%

Covalent attachment of oligodeoxyribonucleotides to amine-modified Si (001) surfaces

Strother

Hamers

Smith

2000

Nucleic Acids Research

279

308

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A recently described reaction for the UV-mediated attachment of alkenes to silicon surfaces is utilized as the basis for the preparation of functionalized silicon surfaces. UV light mediates the reaction of tbutyloxycarbonyl (t-BOC) protected ω-unsaturated aminoalkane (10-aminodec-1-ene) with hydrogenterminated silicon (001). Removal of the t-BOC protecting group yields an aminodecane-modified silicon surface. The resultant amino groups can be coupled to thiol-modified oligodeoxyribonucleotides using a heterobifunctional crosslinker, permitting the preparation of DNA arrays. Two methods for controlling the surface density of oligodeoxyribonucleotides were explored: in the first, binary mixtures of 10-aminodec-1-ene and dodecene were utilized in the initial UV-mediated coupling reaction; a linear relationship was found between the mole fraction of aminodecene and the density of DNA hybridization sites. In the second, only a portion of the t-BOC protecting groups was removed from the surface by limiting the time allowed for the deprotection reaction. The oligodeoxyribonucleotide-modified surfaces were extremely stable and performed well in DNA hybridization assays. These surfaces provide an alternative to gold or glass for surface immobilization of oligonucleotides in DNA arrays as well as a route for the coupling of nucleic acid biomolecular recognition elements to semiconductor materials.

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Modification of the 5′ Terminus of Oligonucleotides for Attachment of Reporter and Conjugate Groups

Thuong

Asseline

2000

CP Nucleic Acid Chemistry

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Reporter and conjugate groups can be added directly to the 5' terminus of oligonucleotides by appropriate modification. Conjugate groups can be used to increase the affinity of complementary strands, induce irreversible modification of target sequences, or enable sequences to recognize and permeate target cell membranes. This overview discusses the 5' modifications that can be used and strategies for the covalent attachment of ligands to the modified oligonucleotides. Step-by-step protocols for attachment of conjugate groups are given elsewhere in the series.

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Immobilization of DNA via oligonucleotides containing an aldehyde or carboxylic acid group at the 5' terminus

Cited by 103 publications

References 33 publications

An improved gel‐based DNA microarray method for detecting single nucleotide mismatch

An improved gel‐based DNA microarray method for detecting single nucleotide mismatch

Covalent attachment of oligodeoxyribonucleotides to amine-modified Si (001) surfaces

Modification of the 5′ Terminus of Oligonucleotides for Attachment of Reporter and Conjugate Groups

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