End labeling is a favored method for applications where an
internal label might interfere with hybridization or sequence specific
protein binding.
5’ or 3’ EndTag™ labeled nucleic acids can be used for
applications such as DNA hybridization, PCR, in situ
hybridization, the binding of capture probes to affinity
matrices, or electrophoretic mobility shift assays (EMSA).
Short oligonucleotides are labeled more efficiently with these
systems than with other methods. In addition, end labeling of
oligonucleotides is an economical alternative to having labels
inserted during synthesis.
Both the 5’ EndTag™ and the 3’ EndTag™ Nucleic Acid Labeling
Systems enable the covalent attachment of a variety of
fluorescent dyes, haptens, or affinity tags to the respective
ends of the nucleic acids using thiol-specific chemistry. Labels
containing thiol-reactive groups (maleimides, iodoacetamides,
etc.) can easily be incorporated.
Labeling time is about 1 hour with very little hands-on time.
3’ EndTag™ Labeling Kit
The 3’ EndTag™ system enables simple and uniform labeling
of blunt, overhanging, or recessed 3’ ends of DNA. 3’ end
labeling is the preferred method if the terminal phosphate at
the 5’ end must be preserved.
Labeling using the 3’ EndTag™ Labeling Kit is achieved in two
steps:
1. Terminal deoxynucleotidyl transferase (TdT) incorporates the
thiol-containing nucleotide (SH-GTP) onto the 3’ end.
2. Thiolated DNA is coupled with a thiol-reactive label.
The 3’ EndTag™ Labeling Kit is designed to perform 20 labeling
reactions of up to 0.5 nmols of 3’ ends (e.g. about 4.2 μg of
a 25 base oligonucleotide) per reaction. Thiol-reactive label is
not included in the kit and should be selected separately.
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