Neuro-2a cells were transiently transfected with Sig-1R EYFP
for 24 h and followed by another 24 h of CQ treatments.
FYDR mice carry a direct repeat recombination substrate for which an HR event can restore the full length Eyfp coding sequence (Figure 3A) (Hendricks et al.
Although these data suggest that low dose-rate radiation did not affect the frequency of HR, it remained formally possible that radiation caused silencing of the Eyfp gene (Suzuki a al.
Spontaneous mitotic homologous recombination at an enhanced yellow fluorescent protein (EYFP) cDNA direct repeat in transgenic mice.
Gene expression was induced with tetracycline (1 mg/L), and cells expressing the EYFP tag were sorted out in a FACS-Diva flow cytometer (BD Biosciences).
The transfection efficiency was evaluated in cells expressing LDLR coupled to the EYFP tag and analyzed by flow cytometry.
There is no spillover of signals between the detectors that measure DiD and EYFP.
Transfection efficiencies were visualized by means of the EYFP tag fused to the LDLR, and any differences were allowed for in the calculations.
In the reporter assay presented here, we used two species derived from green fluorescent protein (GFP), one (enhanced yellow fluorescent protein, (EYFP)) to measure the promotion and enhancement of gene expression, and the other (enhanced cyan fluorescent protein, (ECFP)) to normalize the transfection, and were thus able to measure the fluorescent protein signals simultaneously without any co-factor or substrates.
For reporter plasmids, the luciferase region of the pGL3-Control Vector (Promega, Madison, WI, USA) was replaced with the EYFP fragment of pEYFP-N1 or the ECFP fragment of pECFP-N1 (Clontech, Palo Alto, CA, USA) using NcoI and Xbal sites.
The fluorescence of EYFP was detected with an excitation filter of 485 nm and an emission filter of 545 nm, and that of ECFP was detected with filters of 420 and 486 nm (Perkin Elmer), respectively.
(x) GAL4, UAS-mCD8: GFP larvae were placed on a line separating the two halves of the dish.