Analysis of 3′-amino-tailed tRNA by biotin-streptavidin conjugation. (A) Denaturing 12% PAGE/7M urea gel analysis of conjugation on a Bio-Rad mini-gel apparatus, showing the up-shifted bands of tailed tRNA and the starting band of untailed tRNA. The gel was loaded with ∼70 ng of tRNA in each lane and stained by SYBR gold. In the first lane, no upshifted band was observed with unreacted control (C) tRNA. (B) The tailing frequency of each protocol is calculated from the sum of all up-shifted bands over the amount of both shifted and unshifted bands. These data show that protocol 2 produces the highest level of 3′-amino-tailing. Errors are standard deviation and n = 3.
