Characterization of PicoGreen Reagent and Development of a Fluorescence-Based Solution Assay for Double-Stranded DNA Quantitation

Singer, Victoria L.; Jones, Laurie J.; Yue, Stephen; Haugland, Richard P.

doi:10.1006/abio.1997.2177

Cited by 704 publications

(545 citation statements)

References 25 publications

(11 reference statements)

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“…Samples were also quick frozen on dry ice between purification steps as an additional precaution. DNA quantitation by UV absorbance at 260 nm was complicated by contaminating RNA, so the DNA was quantified instead by a double strand-specific PicoGreen assay method (41). A high dose of UVB light (4000 J/m 2 ) was needed to obtain sufficient signal over background, and under these conditions, the CPD frequency was about one per 500 -1000 bases.…”

Section: Determining Nucleosome Positioning By Hydroxyl Radicalmentioning

confidence: 99%

Rapid Deamination of Cyclobutane Pyrimidine Dimer Photoproducts at TCG Sites in a Translationally and Rotationally Positioned Nucleosome in Vivo

Cannistraro

Pondugula

Song

et al. 2015

Journal of Biological Chemistry

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Background: DNA photoproduct deamination contributes to mutations. Results: Cyclobutane pyrimidine dimers (CPDs) deaminate fastest at TCG sites, and the rate depends on the position of the CPD in a nucleosome determined from hydroxyl radical footprinting. Conclusion: Deamination could explain the high C to T mutation rate at TCG sites, which can be further modulated by nucleosomes. Significance: Sequence context and chromatin structure can modulate UV mutagenesis.

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Section: Determining Nucleosome Positioning By Hydroxyl Radicalmentioning

confidence: 99%

Rapid Deamination of Cyclobutane Pyrimidine Dimer Photoproducts at TCG Sites in a Translationally and Rotationally Positioned Nucleosome in Vivo

Cannistraro

Pondugula

Song

et al. 2015

Journal of Biological Chemistry

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“…The captured nucleic acids were released by electrochemically-cleaving the thiol-gold bond 27 . Desorption of the hybrids was confirmed by an optical assay utilizing PicoGreen, which is a probe that forms a highly fluorescent complex upon specific binding with double-stranded DNA (dsDNA) 28 . We further evaluated the efficacy of the purification process by performing capillary electrophoresis on the samples before and after purification.…”

Section: Resultsmentioning

confidence: 99%

Sequence-Specific Electrical Purification of Nucleic Acids with Nanoporous Gold Electrodes

et al. 2016

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Nucleic acid-based biosensors have enabled rapid and sensitive detection of pathogenic targets; however, these devices often require purified nucleic acids for analysis since the constituents of complex biological fluids adversely affect sensor performance. This purification step is typically performed outside the device, thereby increasing sample-to-answer time and introducing contaminants. We report a novel approach using a multifunctional matrix, nanoporous gold (npAu), which enables both detection of specific target sequences in a complex biological sample and their subsequent purification. The np-Au electrodes modified with 26-mer DNA probes (via thiolgold chemistry) enabled sensitive detection and capture of complementary DNA targets in the presence of complex media (fetal bovine serum) and other interfering DNA fragments in the range 50 to 1500 base pairs. Upon capture, the non-complementary DNA fragments and serum constituents of varying sizes were washed away. Finally, the surface-bound DNA-DNA hybrids were released by electrochemically cleaving the thiol-gold linkage and the hybrids were iontophoretically eluted from the nanoporous matrix. The optical and electrophoretic characterization of the analytes before and after the detection-purification process revealed that low target DNA concentrations (80 pg/µL) can be successfully detected in complex biological fluids and subsequently released to yield pure hybrids free of polydisperse digested DNA fragments and serum biomolecules. Taken together, this multifunctional platform is expected to enable seamless integration of detection and purification of nucleic acid biomarkers of pathogens and diseases in miniaturized diagnostic devices.

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“…Picogreen works as a DNA identifier 16 as it emits characteristic green emission only when it intercalates within the DNA double helix structure. Figure 4 shows the photoluminescence (PL) spectra of several DNA-CTMA films with different thickness stained with Picogreen dye.…”

Section: Report Documentation Pagementioning

confidence: 99%

Molecular Beam Deposition of DNA Nanometer Films

Hagen

Spaeth

et al. 2006

Nano Lett.

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The development of novel photonic devices which incorporate biological materials is strongly tied to the development of thin film forming processes. Solution-based ("wet") processes when used with biomaterials in device fabrication suffer from dissolution of underlying layers, incompatibility with clean environment, inconsistent film properties, etc. We have investigated ultra-high-vacuum molecular beam deposition of surfactant-modified deoxyribonucleic acid (DNA). We have obtained effective deposition rates of ∼0.1−1 Å/s, enabling reproducible and controllable deposition of nanometer-scale films.Interest is continuing to increase 1-5 rapidly in the optical and electronic properties of DNA and other biopolymers and in related device applications. Usually, reports on these properties are either based on relatively thick films obtained by "wet" processes (such as spin-coating) techniques 6 or based on heroic efforts with single molecules. 7,8 In this paper, we report on the properties of nanometer-scale surfactantmodified DNA thin films formed by molecular beam deposition [9][10][11] (MBD). In general, polymers, unlike small molecule organic materials, do not usually lend themselves to MBD since their high molecular weight results in very low vapor pressures (and negligible deposition) up to their decomposition temperature. However, we have found that several types of complexed DNA can be deposited by MBD with subnanometer thin film control, thus opening the door to its incorporation in nanoscale devices. MBD is a thermal evaporation technique widely utilized in the fabrication of photonic devices based on compound semiconductors, where it is commonly known as molecular beam epitaxy (MBE) since the thin films have an epitaxial relationship to the substrate on which they are grown. This deposition technique takes place under high vacuum conditions, which allows the formation of a molecular beam (with a minimum of scattering) and results in deposition rates of the order of 1 Å/s for atomic or small molecule materials. The small furnace (or effusion cell) that holds the material to be evaporated is connected to the high vacuum chamber. The deposition rate is controllable with high precision over a wide range by adjusting the temperature of the effusion cell containing the material. An MBD chamber with several cells can be utilized for the sequential deposition of multiple materials for the formation of complex device structures. This in situ "dry" process in a high vacuum environment prevents the generation of defects by contamination of the materials or by particulate deposition from the environment, both of which can occur during wet processing.We have previously reported 12 on the first use of spincoated DNA complex films as electron "blocking" layers (EBL) in organic light-emitting devices (OLEDs), showing significant enhancement in both luminance and device efficiency over conventional device structures. In this paper, we report on the properties of nanometer-thin DNA-based films by the MBD technique.We...

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Characterization of PicoGreen Reagent and Development of a Fluorescence-Based Solution Assay for Double-Stranded DNA Quantitation

Cited by 704 publications

References 25 publications

Rapid Deamination of Cyclobutane Pyrimidine Dimer Photoproducts at TCG Sites in a Translationally and Rotationally Positioned Nucleosome in Vivo

Rapid Deamination of Cyclobutane Pyrimidine Dimer Photoproducts at TCG Sites in a Translationally and Rotationally Positioned Nucleosome in Vivo

Sequence-Specific Electrical Purification of Nucleic Acids with Nanoporous Gold Electrodes

Molecular Beam Deposition of DNA Nanometer Films

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