Supplements
Analysis of Spo11-Myc18 functionality by random spore analysis.
Random spore analysis was performed as described previously (Loidl et al. 1998). Briefly, single colonies of strains carrying his4X, his4B heteroalleles were picked, grown in 5ml of YPD followed by YPA. Upon transfer to sporulation medium an aliquot was removed, counted and plated on SM-HIS to determine the frequency of His+ cells that arose during mitosis. The mitotic background of wild type (YFK47) was 2/104 and that of Spo11-Myc18 (YFK2075) 16,7/104 His+ cells at t=0. Cells were sporulated at 16°C, 30°C and 34°C for two days. Asci were digested with zymolyase and sonicated until more than 95% of the spores were single. No intact vegetative cells were seen at that point. Spores were then counted using a haemocytometer and 150.000 spores were plated directly on SM-HIS, while a total of 5.000 spores were plated on YPD plates and later replica plated to SM-HIS. Supplementary Table 1S has the results of this analysis.
Supplementary Table 1S
temp (°C) / Spo11 / Spo11-Myc18 / Spo11 / Spo11-Myc18 / Spo11 / Spo11-Myc18Sporulation / Spore viability in % / His+/104 viable spores *
16 / 81,5 / 83 / 97 / 96 / 99/86 / 80/94
30 / 93 / 94 / 100 / 100 / 114/125 / 176/178
34 / 93,5 / 94 / 97 / 100 / 57/88 / 53/71
* The first number represents the His+ colonies when spores were plated directly on SM-HIS, the number after the slash represents the His+ colonies scored after replica plating to SM-HIS of colonies first grown on YPD.
References
Loidl, J., Klein, F., and Engebrecht, J. 1998. Genetic and morphological approaches for the analysis of meiotic chromosomes in yeast. In Nuclear structure and function (ed. M. Berrios), pp. 257-285. Academic Press., San Diego.