Fig.2.

Scheme representing the use of chronological life span to monitor age-dependent mutation frequency. A. Detection of point mutations+frameshifts+complex mutational events: 2 10⁷ cells are plated on SDC-ARG+canavanine every 2 days. Can^r mutation frequency is calculated based on the number of viable cells as measure by CFUs assay. B. Detection of point mutations: 10⁸ cells are plated on SDC-TRP every 2 days. Trp+colonies growth is caused by reversion of an amber stop codon (corresponding to residue 135 of the coding sequence). C. Detection of GCRs: 10⁸ cells are plated on SDCARG+canavanine+5FOA every 2 days. Can^r 5FOA^r cells growth is mostly caused by loss of a large region of DNA that contains both the CAN1 and the URA3 genes (Chen and Kolodner, 1999).

D. Detection of frameshift mutaions: 10⁸ cells are plated starting from a YPD overnight inoculum on SDC-LYS. Lys+ colonies are scored every day and their growth is due mostly to small insertions (2 or 2+3n) or small deletions (1 or 1+3n). The frequency of frameshifts is calculated every day by dividing the cumulative number of colonies on the plate by the number of viable cells.