Figure 5
From: Auxin-inducible protein depletion system in fission yeast
Application of the i -AID system to other replication factors. (A) Derivatives of the P adh15 -skp1-AtTIR1-NLS strain carrying mcm4-aid (HM2473), mcm10-aid (HM2550), cdc20-aid (HM2551), orc2-aid (HM2572), pol1-aid (HM2552), orc6-aid (HM2575) and cdc45-aid (HM2578) were spotted on EMM plates containing auxin (0.5 mM) and incubated at 25°C. (B) Flow cytometry analysis of the derivatives shown in (A). Each strain was incubated in the presence of auxin (0.5 mM) at 25°C and the DNA contents were analyzed. (C) Immunoblotting analysis of Mcm10-aid. HM2550 P adh15 -skp1-AtTIR1-NLS mcm10-aid cells incubated in the presence of auxin (0.5 mM) at 25°C and cell extracts prepared at the indicated time points were subjected to immunoblotting with anti-Mcm10 and anti-TAT1 antibodies. The 0 h sample was diluted to 50%, 25% and 12.5% with SDS sample buffer, and used for comparison of the protein amounts. Non-specific protein bands are indicated by asterisks (*).