FIG. 3.
Construction of the cdc20ΔN-term mutant. (A) Expected genomic structure of the cdc20 and leu1 loci following integration of pJK148-cdc20C1. The solid and cross-hatched bars indicate the regions of cdc20 corresponding to the N terminus and the C terminus, respectively. P indicates the location of the PstI restriction sites used for the Southern blot analysis. (B) Southern blot of genomic DNA prepared from the diploid Δcdc20/cdc20+ strain (lane 3) and from two independent cdc20ΔN-term isolates (lanes 1 and 2), probed with a PCR fragment corresponding to the C-terminal half encoded by cdc20. (C) Western blot of a protein extract prepared from wild-type cells (lane 1) and cdc20ΔN-term cells (lane 2), using anti-HA monoclonal antibodies. The apparent molecular mass of 3HACdc20C1p is approximately 122 kDa.