Figure 5.

Hat1-TAP is recruited to early and late origins at the time of firing. Samples were taken at indicated times and processed for chromatin immunoprecipitation and input controls. DNA was either labeled with ³²P (panels A and C) or visualized by ethidium bromide (panels B and D). (A) Binding of Hat1-TAP and Cdc45-TAP to ARS305 and to R11 control sequence at the time of origin firing and later in S-phase. Strains were HAT1-TAP (BSY679) and CDC45-TAP (BSY680). One representative experiment with input and immunoprecipitate and the percentage of precipitated DNA is shown. (B) Recruitment of Hat1p to ARS1 is coincident with Cdc45p and dependent on functional ORC. Strains used for chromatin immunoprecipitation are HAT1-TAP, CDC45-TAP, and HAT1-TAP orc2-1 (BSY699). Strains were held in a-factor at 36°C to inactivate orc2-1 and then released at 23°C. (C) Recruitment of Hat1p (Hat1-TAP) to ARS1412 late replication origin and comparison with R11 sequence. (D) Recruitment of Hat1p to ARS1 is affected by the cdc7-1 allele. Strains used for chromatin immunoprecipitation are HAT1-TAP, CDC45-TAP, and HAT1-TAP cdc7-1 (BSY734).