End labeled oligonucleotides

Gel electrophoresis of P-end-labelled oligonucleotides irradiated with visible light in the presence of RufTAP)] showed that the principal photochemical product is a less electrophoretically-mobile species [96]. This is consistent with the formation of a photo-adduct and it is clear that the yield of this reaction is... 

Figure 5.48 Molecular structures of two of the photoactive pyrene end-labeled oligonucleotides reported by Reese and Fox [80]...

Figure 5.49 Time course for the limiting current at gold modified with the oligonucleotide 2/4 in methylviologen (MV+2) solution (0.1 mM in 0.1 NaCl) at an applied potential of 0 mV (vs. Ag/AgCl). The inset shows the time domain in which the electrode is alternatively exposed to and shielded from 346 nm illumination the arrows indicate when the shutter is open or closed. From R. S. Reese and M. A. Fox, Spectral and cyclic voltammetric characterization of self-assembled monolayers on gold of pyrene end-labeled oligonucleotide duplexes, Can.. Chem., 77,1077-1084 (1999), with permission from The National Research Council of Canada...

The highly radioactive end-labeled oligonucleotides should not be used for more than 1 week after preparation because they are subject to radiochemical degradation. Labeled probes not used immediately are stored at — 20°. We routinely prepare several groups of slot blots and perform the hybridization to oligonucleotide probes on the day they are labeled. 

G 4.O-NH2 to obtain a thin fiim. Single-stranded 3 -biotin end-labeled oligonucleotide was immobilized onto the film to obtain a stable recognition layer through biotin-avidin combination to detect complementary target. 

Figure 1.72 Cystamine may be used to label phosphate groups, such as at the 5 -end of oligonucleotides, via a carbodiimide reaction using EDC. The resultant phosphoramidate linkage is a common way to modify oligonucleotides at the 5 -end.

In order to facilitate analysis of FeBABE produced fragments, the prey protein or biomolecule is labeled at one end with a tag that can be detected after electrophoresis, usually in a transfer blot. The tag can be a fusion tag, such as 6X His, or any other group that can be targeted with an antibody and detected. Alternatively, radiolabels and fluorescent labels have been used with prey molecules, including the use of end-labeled DNA to study where DNA binding proteins dock onto the oligonucleotide sequence. 

The donor-acceptor distance may not be unique, especially when the donor and acceptor are linked by a flexible chain (e.g. end-labeled oligomers of polymethylene or polyethylene oxide, oligopeptides, oligonucleotides, polymer chains). 

Assay for human immunodeficiency virus type 1 (HIV-1) proviral DNA in peripheral blood monuclear cells can be performed by PCR followed by detection of PCR products by electrochemiluminescence-labeled oligonucleotide probe [Tris-bipyridine ruthenium (II) complex]. Since one of the PCR primers is biotin-labeled at the 5 end, facile capture of the PCR product-probe complex can be accomplished on streptavidin-conjugated magnetic particles, prior to analysis in an electrochemiluminescence analyzer (S3). 

Fig. 2.11. In (a), 3 -end labelling is achieved using deoxynucleotidyl transferase and an ff-[32P] ribonucleotide triphosphate followed by elimination of the ribonucleotide residues. In (b), 5 -end labelling is carried out using polynucleotide kinase and y-[32P]ATP. Partial digestion with spleen phosphodiesterase removes nucleotides sequentially from the 5 -end of the oligonucleotide giving a mixed population of partially degraded molecules each labelled at the 3 -end. Venom phosphodiesterase removes nucleotides from the 3 -end similarly yielding a mixed population of shortened fragments. The products of the reaction are resolved by two-dimensional electrophoresis-homochromatography and the sequence deduced by the characteristic...

Guerra CE. Analysis of oligonucleotide microarrays by 3 end labeling using fluorescent nucleotides and terminal transferase. Biotechniques 2006 41 53-56. 

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