Add internal markers or label at the ends
There are two ways to label DNA with enzymes: at the ends or internally along the molecule. Detectable markers such as fluorophores or modified nucleotides can be introduced by the addition of 5’ and 3’ groups through enzyme action, as well as by introducing internal markers through direct incorporation or post-labeling techniques with a suitable enzyme.
Enzyme strategies for adding labels to nucleic acids
Labeling by the action of an enzyme is required for many studies aiming to elucidate the functions and dynamics of nucleic acids in vitro and in cells. For every site where labeled nucleotides can be added to DNA or RNA, there is a suitable enzyme that can catalyze the addition.
| Enzyme | Activity | Labeling action | ||||
|---|---|---|---|---|---|---|
| End labeling |
Direct incorporation |
By random priming |
Gap label | Nick translation |
||
| P7060L Klenow Fragment |
DNA polymerase with 5’→3’ |
✘ No | ✔ Yes | ✔ Yes | ✘ No | ✘ No |
| P7010 Klenow (3’→5’ exo-) |
DNA polymerase with 5’→3’ synthesis activities |
✔ Yes (3’ dsDNA) | ✔ Yes | ✔ Yes | ✘ No | ✘ No |
| P7260L T7 DNA polymerase Coming soon |
DNA polymerase with 5’→3’ synthesis and 3’→5’ ssDNA and dsDNA exonuclease activity. [The double-stranded DNA exonuclease activity requires the presenceof thioredoxin.] |
✔ Yes (5’ dsDNA) | ✔ Yes | ✘ No | ✔ Yes | ✘ No |
| P7050L DNA polymerase I |
DNA polymerase with 5’→3’ synthesis, 5’→3’ and 3’→5’ exonuclease activities |
✘ No | ✘ No | ✘ No | ✘ No | ✔ Yes |
| RP810 TaqNova Stoffel DNA Polymerase BLIRT |
Stoffel Fragment derivatives of Taq DNA polymerase that is 5’→3’ exonuclease deficient |
✘ No | ✔ Yes | ✘ No | ✘ No | ✘ No |
| P7620L TaqIT DNA polymerase |
TaqIT is 5’→3’ exonuclease deficient derivative of Taq DNA polymerase (Stoffel) |
✘ No | ✔ Yes | ✘ No | ✘ No | ✘ No |
| P7070L Terminal deoxynucleotidyl transferase (TdT) Coming soon |
Catalyzes the addition of dNTPs to the 3' hydroxyl terminus of DNA molecules; extends blunt, 5’-overhanging, ssDNA and RNA |
✔ Yes (3’ ssDNA) | ✘ No | ✘ No | ✘ No | ✘ No |
| P7180L T7 RNA polymerase Coming soon |
Synthesizes RNA in 5’→3’ direction off DNA template |
✘ No | ✔ Yes (RNA) | ✘ No | ✘ No | ✘ No |
| P7460L Poly(A) polymerase |
Catalyzes the addition of AMP from ATP to the 3’ hydroxyl of RNA for poly(A) tailed RNA |
✔ Yes (3’ RNA) | ✘ No | ✘ No | ✘ No | ✘ No |
| L6050L T4 RNA ligase 1 Coming soon |
Ligates RNA and ssDNA molecules |
3’ (RNA and ssDNA) | ✘ No | ✘ No | ✘ No | ✘ No |
| Y9040L T4 polynucleotide kinase (PNK) |
Catalyzes the transfer of the γ-phosphate from ATP to the 5´-terminus of polynucleotides or to mononucleotides bearing a 5´-hydroxyl group |
5’ (ssDNA, dsDNA and RNA) |
✘ No | ✘ No | ✘ No | ✘ No |
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