Localization of exogenous Cyclin E in pre-MBT and MBT Xenopus laevis embryos. One cell of 2-cell embryo was microinjected with in vitro transcribed Myc6−GFP-Cyclin E RNA, collected at indicated time points, and the translated protein detected in fixed and stained embryos. For immunofluorescence analysis of Cyclin E localization, embryos were collected at 4 hpf, pre-MBT (a-c) or at 6 hpf, MBT (d-f). (a, d) Embryos were fixed and stained with an antibody against the Myc6 tag (αMyc) followed by an Alexa488 conjugated secondary antibody. (b, e) Embryos were counterstained with DAPI to visualize the nuclei. (c, f). Merged image of the Alexa488 and DAPI. White arrowheads in d-f indicate nuclei. Embryos were viewed with a 10X objective on a Zeiss LSM 510 confocal microscope equipped with a META detector, and analyzed using LSM510 Image Acquisition software. Scale bars are 100 μM. At least 20 embryos were injected in at least 3 separate experiments, with at least 5 embryos fixed per timepoint for analysis.