Bulky dyes may block the ribosome during translation and we have seen lower translational efficiencies with more substitutions. We recommend not to fully substitute, but only substitute 10-50%. For our catalog products, we use a 25% substitution of the dye-labeled nucleotide. Please contact us for any additional information.
TriLink offers a variety of modified rNTPs suitable for in vitro transcription. We have assessed transcription efficiency based on final product formation of a 1.9 kb transcript with the following triphosphates at 100% substitution using T7 polymerase.
+++ = Greater than 75% efficiency compared to unmodified NTP
++ = Between 25-75% efficiency compared to unmodified NTP
+ = Less than 25% efficiency compared to unmodified NTP
n/a = No significant product formed with 100% substitution
In all cases, transcription was carried out at 37°C.
You can confirm translation by western blot or enzyme-linked immunosorbent assay (ELISA).