fragment. The level of imprinting was determined by the amount of the mat1 fragments that were broken during the purification process. The intensity of signals was measured using a BioRad Phosphorimager. Two independent measurements were per- formed for each strain and the average was given in the figures. 2D-gel analysis of replication intermediates 2D gels were performed as described in [38], except the first- and second-dimension gels contained 0.5% and 1.2% agarose, respectively. The replication intermediates were digested with the restriction enzymes given in Figure 2C, Figure 5E, Figure 7, and Figure 8C. The intensity of the replication signals was determined using a BioRad Phosphorimager. The pause signal level was given relative to the intensity of the ascending Y-arc, as well as to the intensity of the wild-type pause signal. The average measurement from two gels was given for each strain. High-resolution Southern blot analysis of replication intermediates Yeast chromosomal DNA was purified as described in [39]. The replication intermediates were digested with the restriction enzymes given in Figure 5A. Replication intermediates were separated from non-replicating DNA by BND-cellulose chroma- tography [40] and separated on a 4% denaturing polyacrylamide gel. After electrophoresis, the DNA was transferred to a GeneScreen membrane (NEN) by electro-blotting, and probed with strand-specific probes that had been generated as described in [41]. Precise mapping of the position of the imprint Chromosomal DNA was purified as described before [10]. The DNA were digested with SspI and precipitated overnight with References 1. Tajbakhsh S (2008) Stem cell identity and template DNA strand segregation. Curr Opin Cell Biol 20: 716–722. 2. Miyata H, Miyata M (1981) Mode of conjugation in homotallic cells of Schizosaccharomyces pombe. J Gen Appl Microbiol 27: 365–371. 3. 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