Abnormal transition from conjugal development to the first vegetative cell division in heterozygous C3ΔRFB/B progeny. ( A ) Normal kinetics for pair formation and separation in C3ΔRFB mutant progeny. Microscopic analysis of pair formation and separation in wild-type crosses [SB210 × TX614(1)], crosses between wild-type and mutant strains [SB210 × TX607(1), SB210 × TX610(11), SB210 × TX611(1)] and cross between two mutant strains [TX607(1) × TX610(11)]. ( B ) Cytological examination of crosses between wild-type strain SB210 and wild-type C3 rDNA transformant, TX614(1) (WT × WT),or SB210 and C3ΔRFB transformant, TX607(1) (WT × Mutant) with the DNA staining dye apofluor. The vast majority of cells displayed normal progression through development (data not shown), with a small percentage of mutant mating partners containing extra micronuclei/pronuclei prior to genetic exchange (compare micrographs 1 and 6). Panels 2–5 (WT × WT) and 7 ×10 (WT × Mutant) depict representative cells in exconjugant mating populations (24 h mating followed by 7–9 h re-feeding). ( C ) Cartoon depicting the progression of mating cells during development (0–24 h) and the fate of exconjugants after re-feeding at 24 h for (WT × WT), and (WT × mutant) or (mutant × mutant) crosses. See text for details. ( D ) Quantification of the number of ‘abnormal cells’ for different mating 7–9 h after re-feeding (see text for details). ( E ) Progeny of crosses between TX611(1) and A* mating type III obtained by single pair isolation after 23–25 h and subsequent propagation for 5–10 passages. Note the appearance of extra nuclei in the mutant (micrographs 2–4) compared with wild-type (micrograph 1). Staining: apofluor (micrographs 1–3), DAPI (micrograph 4).