DNA, hybridization

The DNA hybridization conditions may vary depending on the membrane used, and suppliers will often give details of optimal conditions to use with their membrane. 

The following conditions work well with Hybond N and N (Amersham) membranes using a prehybridization/hybridization buffer made according to the following recipe. For 50 ml of buffer, combine 40 ml sterile distilled 

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Fig. 3.46. " Sn image of Sn-labeled DNA hybridized to six oligonucleotide spots three complimentary and three non-comple-mentary.

If DNA from two different species are mixed, denatured, and allowed to cool slowly so that reannealing can occur, artificial hybrid duplexes may form, provided the DNA from one species is similar in nucleotide sequence to the DNA of the other. The degree of hybridization is a measure of the sequence similarity or relatedness between the two species. Depending on the conditions of the experiment, about 25% of the DNA from a human forms hybrids with mouse DNA, implying that some of the nucleotide sequences (genes) in humans are very similar to those in mice. Mixed RNA DNA hybrids can be created in vitro if single-stranded DNA is allowed to anneal with RNA copies of itself, such as those formed when genes are transcribed into mRNA molecules. 

Verhaegen, M., and Christopoulos, T. K. (2002). Recombinant Gaussia luciferase. Overexpression, purification, and analytical application of a bioluminescent reporter for DNA hybridization. Anal. Chem. 74 4378-4385. 

FIGURE 6-14 DNA hybridization biosensors detection of DNA sequences from the E. coli pathogen. Chronopotentiometric response of the redox indicator upon increasing the target concentration in 1.0 pg/ml steps (a-c), in connection with a 2 min hybridization time. (Reproduced with permission from reference 46.)... 

Current analytical methods have difficulty detecting picogram levels of nucleic acids, particularly when high levels of other biopolymers (e.g., proteins) are present. The most widely used assay method employed by the pharmaceutical industry involves a nick translation DNA hybridization method (1). This assay offers high sensitivity and selectivity but has a number of drawbacks. 

TABLE 10 Results of Fluorescent and Gravimetric Measurements of DNA Hybridization Against a Reference Containing only LB-Immobilized Single-Stranded DNA/Amine Sandwich... 

The use of DNA hybridization just described opens up a novel, uncomplicated, yet powerful strategy for supramolecular synthesis Many different devices are connected to a distinct sequence codon and are subsequently organized on a suitable template strand. The utilization of appropriate nucleic acid scaffolds should even allow the fabrication of highly complex supramolecular structures by means of a modular construction kit. For approximately 20 years, the work of Seeman and coworkers [8,27] have been engaged in the rational construction of ID, 2D, and 3D DNA frameworks. They use branched DNA... 

Bifunctional adamantyl, as a hydrophobic central core, can be used to construct peptidic scaffolding [151], as shown in Fig. 27. This is the reason why adamantane is considered one of the best MBBs. This may be considered an effective and practical strategy to substitute different amino acids or DNA segments on the adamantane core (Fig. 28). In other words, one may exploit nucleic acid (DNA or RNA) sequences as linkers and DNA hybridization (DNA probe) to attach to these modules with an adamantane core. Thus a DNA-adamantane-amino acid nanostructure may be produced. 

Fig. 6.5 Schematic representation of a bioelectronic protocol for detection of DNA hybridization (A) binding of the target to magnetic beads (B) hybridization with CdS-labeled probe (C) dissolution of CdS tag (D) potentiometric stripping detection at a mercury-film electrode. (Reprinted from [136], Copyright 2009, with permission from Elsevier)...

DNA hybridization based on cadmium sulfide nanoparticle tags. Electrochem Commun 4 ... 

Voordouw G, JK Voordouw, RR Karkhoff-Schweiser, PM Eedorak, DWS Westlake (1991) Reverse sample genome probing, a new technique for identification of bacteria in environmental samples by DNA hybridization, and its application to the identification of sulfate-reducing bacteria in oil field samples. Appl Environ Microbiol 57 3070-3078. 

Several laboratory tests are available to aid in the diagnosis of gonorrhea and include gram-stained smears, culture, or the DNA hybridization probe. 

Tests used to diagnose C. trachomatis include culture, the enzyme immunoassay, the DNA hybridization probe, or the... 

Grozema FC, Siebbeles LDA, Gelinck GH, Warman JM (2005) The Opto-Electronic Properties of Isolated Phenylenevinylene Molecular Wires. 257 135-164 Guiseppi-Elie A, Lingerfelt L (2005) Impedimetric Detection of DNA Hybridization Towards Near-Patient DNA Diagnostics. 260 161-186 Guo Y, see Coleman AW (2007) 277 31-88... 

Collins, M. L., et al. (1995). Preparation and characterization of RNA standards for use in quantitative branched-DNA hybridization assays. Anal. Biochem. 226,120-129. 

Valis L, Mayer-Enthart E, Wagenknecht H-A (2006) 8-(Pyren-l-yl)-2 -deoxyguanosine as an optical probe for DNA hybridization. Bioorg Med Chem Lett 16 3184-3187... 

This enzyme is associated with the virions of RNA tumor viruses such as the Rous sarcoma virus (RSV). The enzyme has remarkable enzymatic activity in that it can catalyze several seemingly diverse steps in the synthesis of double-stranded DNA from the single-stranded RNA viral genome. The enzyme uses a tRNA for tryp-tophan as a primer to make a copy of DNA that is complementary to the viral RNA. The resulting RNA-DNA hybrid is converted to a double-stranded DNA molecule by ribon-uclease (RNase)H and DNA-dependent DNA polymerase activities that are intrinsic to reverse transcriptase. 

Griffiths B.S., Diaz-Ravina M., Ritz K., McNicol J.W., Ebblewhite N., Baath E. Community DNA hybridization and %G+C profiles of microbial communities from heavy metal polluted soils. FEMS Microbiol. Ecol. 1997 24 103-112. 

Freeman WM et al. Fundamentals of DNA hybridization arrays for gene expression analysis. BioTechniques 2000 29 1042-1055. 

Lanthanides also have potential as DEFRET energy donors. Selvin et al. have reported the use of carbostyril-124 complexes (53) with europium and terbium as sensitizers for cyanine dyes (e.g., (54)) in a variety of immunoassays and DNA hybridization assays.138-140 The advantage of this is that the long lifetime of the lanthanide excited state means than it can transfer its excitation energy to the acceptor over a long distance (up to 100 A) sensitized emission from the acceptor, which occurs at a wavelength where there is minimal interference from residual lanthanide emission, is then measured. 

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