- Gently swirl the dish in a clockwise and anticlockwise direction
so that the transfection mix covers the surface of the cells. Place
the dish in the CO 2 incubator at 37C for 1 h. - Wash the cells with 1 mL of complete DMEM and add 1 mL of
precooled Triton X 0.01% solution containing 40 μMof
cyclooctyne dye Cy3 DBCO. Incubate at 4C for 12 min
and wash three times with 1 mL of 1PBS. - Supplement the cells with 1 mL of freshly prepared DMEM
containing 40μM of Cy3 DBCO. Place the dish immediately in
the incubator at 37C for 30 min.
Fig. 4 Imaging cellular RNA transcription using AMUTP under copper-free
SPAAC reaction conditions. HeLa cells were transfected with 1 mM of AMUTP
for 1 h. The cells were fixed and stained by SPAAC reaction using Cy3 DBCO. This
figure has been reproduced by permission of Nucleic Acids Research: Oxford
Journals [24]
Imaging Newly Transcribed RNA in Cells 367