Figure 7.

Cen1 protein levels decrease when CEN1 is deleted. (A) Southern blot analysis of the CEN1 locus. The fragment used for transformation extends from the SacI site to the PstI site with the NEO2 cassette replacing CEN1. Total Tetrahymena DNA from wild-type cells (lane 1), the micronuclear transformant (lane 2), and progeny from the knockout heterokaryon strains rescued by CEN1 transformed at a second genomic location (lane 3) was digested with NsiI and PstI. The sizes of select marker bands (M) are noted in kilobase pairs. (B) Western blot analysis of progeny from the cen1Δ knockout heterokaryon strains UCB8 and UCB9 (cen1Δ) and of the progeny from wild-type strains B2086 and CU428 (CEN1). Cen1 is visualized with the Cen1 polyclonal antibody, and a mAb to α-tubulin is used as a loading control. V, vegetative growth; S, starvation; numbers correspond to hours after parental cells were mixed. Marker bands are noted, and sizes are in kilodaltons.