Figure 3.

Together the Mcm2 and Mcm3 NLSs are sufficient to direct the nuclear localization of a heterologous protein. Overnight cultures of YJL310 containing URA3-marked centromeric plasmids pAR109 (pGAL-SV40NLS-GFP₃), pAR110 (pGAL-NLS2-GFP₃), pAR101 (pGAL-NLS3-GFP₃), pAR113 (pGAL-NLS2-NLS3-GFP₃), pAR126 (pGAL-nls2-NLS3-GFP₃), or pAR127 (pGAL-NLS2-nls3-GFP₃) and growing in SRaf-Ura medium were shifted to YEPRaf medium for 90 min before splitting each culture in two and adding α-factor to one-half and nocodazole to the other. One hour later, as the cultures were approaching a complete arrest, galactose was added to induce synthesis of the GFP₃ fusion proteins. After 2 h of induction, cells were examined by fluorescence microscopy.