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[TM] 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[TM] and the 3' EndTag[TM] 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[TM] Labeling Kit
The 3' EndTag[TM] 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[TM] 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.