Figure 1.

Long-term propagation of unc-46 mdf-1 such-4; dog-1 homozygotes for 470 generations results in significant fitness recovery. (A) A schematic representation of the long-term propagation experiment. First, we generated a P0 strain of the following genotype: unc-46(e177) mdf-1(gk2) +/+ + nT1[let-X]; dog-1(gk10)/dog-1(gk10). Since mdf-1(gk2) results in lethality, gk2 is kept balanced over nT1 which is a reciprocal translocation between Chromosomes IV (depicted in red) and V (depicted in blue), and serves as an effective recombination suppressor along the translocated portions of each chromosome. dog-1(gk10) on Chromosome I is depicted in brown. We picked F1 unc-46 mdf-1; dog-1 homozygotes (n = 40) and plated them individually. We isolated a single plate containing fertile worms that survived beyond 3 generations as a suppressor candidate and named it such-4.⁹ The rest of the plates had no surviving progeny after 3 generations (depicted in light gray). We backcrossed one worm from F4 unc-46 mdf-1 such-4; dog-1 to N2 in order to remove dog-1(gk10) and thus avoid further accumulation of dog-1 induced mutations (depicted with light green). A second clone from F4 unc-46 mdf-1 such-4; dog-1 was maintained at 20°C for 470 generations to allow further accumulation of mutations. The worms were frozen for long term storage at the following points: F170 (depicted in dark green), F270 (depicted in purple) and F470 (depicted in blue). (B) Quantification of fitness, measured as the percent of fertile hermaphrodite progeny. Error bars represent SEM (n = 8 trials for each strain and 10 worms per trial). Please note that our initial analysis revealed that F2 unc-46(e177) mdf-1(gk2); dog-1 did not differ significantly from F2 unc-46(e177) mdf-1(gk2) homozygotes (t-test p = 0.845) and F4 unc-46(e177) mdf-1(gk2) such-4(h2168); dog-1 homozygotes did not differ significantly from F4 unc-46(e177) mdf-1(gk2) such-4(h2168) homozygotes (t-test p = 0.357). For that reason we performed multiple trials using F2 unc-46(e177) mdf-1(gk2) (depicted in white) and F4 unc-46(e177) mdf-1(gk2) such-4(h2168) (depicted in light green) as controls.