Figure 4.

Nuclear run-on analysis of nocturnin transcript initiation. Eyecups were prepared in the light and cultured in cyclic light or constant darkness. At the times indicated, retinas were dissected from the eyecups in light (cyclic light samples) or under infrared light (constant dark samples). Thirty retinas were pooled for each time point and nuclei were isolated. Run-on transcriptions were performed with [α-³²P]UTP, using 1 × 10⁷ nuclei per reaction and the resulting labeled transcripts were purified and hybridized to slot-blotted cDNA of vector alone, β-actin, and nocturnin. Filters were exposed to x-ray film and were quantitated by PhosphorImager analysis. The rectangular bars above the graph represent the experimental conditions as described in Fig. 3. The first two samples are from cyclic light on the first day in culture and the last three samples are from constant dark conditions on the second and third day in culture.