Yeast Strain Construction for SGA Analysis

The G1-RNH202-TAP (this study), S-, and G2-RNH202-TAP alleles^{8 (link)} were crossed to the haploid background strain (Y8205, Source C. Boone) for the SGA query strain construction. Selection of diploids, sporulation and tetrad analysis generated the four query strains used in SGA analysis. Cell cycle restricted protein expression of Rnh202 was confirmed by arrest and release experiment and western blot analysis. The selectable markers for SGA analysis were verified by PCR (oMT86/oMT91 for can1Δ::STE2pr-Sp_his5, oMT89/oMT90 for lyp1Δ::STE3pr-LEU2) and replica-plating on YPD + (50 μg/ml canavanine, 50 μg/ml thialysine). The yeast strains are listed in Supplementary Data 2.

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Schindler N., Tonn M., Kellner V., Fung J.J., Lockhart A., Vydzhak O., Juretschke T., Möckel S., Beli P., Khmelinskii A, & Luke B. (2023). Genetic requirements for repair of lesions caused by single genomic ribonucleotides in S phase. Nature Communications, 14, 1227.

Publication 2023

Canavanine Cell cycle Diploids Protein Strains Thialysine Western blot analysis Yeast

Corresponding Organization : Johannes Gutenberg University Mainz

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Variable analysis

independent variables

G1-RNH202-TAP (this study)
G2-RNH202-TAP

dependent variables

Cell cycle restricted protein expression of Rnh202

control variables

Haploid background strain (Y8205, Source C. Boone)

controls

Positive control: Arrest and release experiment and western blot analysis to confirm cell cycle restricted protein expression of Rnh202
Negative control: Not explicitly mentioned

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