Donnelly Centre, University of Toronto, Toronto, ON, Canada.
Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada.
Mol Syst Biol. 2018 May 28;14(5):e7985. doi: 10.15252/msb.20177985.
Condition-dependent genetic interactions can reveal functional relationships between genes that are not evident under standard culture conditions. State-of-the-art yeast genetic interaction mapping, which relies on robotic manipulation of arrays of double-mutant strains, does not scale readily to multi-condition studies. Here, we describe barcode fusion genetics to map genetic interactions (BFG-GI), by which double-mutant strains generated via "party" mating can also be monitored for growth to detect genetic interactions. By using site-specific recombination to fuse two DNA barcodes, each representing a specific gene deletion, BFG-GI enables multiplexed quantitative tracking of double mutants via next-generation sequencing. We applied BFG-GI to a matrix of DNA repair genes under nine different conditions, including methyl methanesulfonate (MMS), 4-nitroquinoline 1-oxide (4NQO), bleomycin, zeocin, and three other DNA-damaging environments. BFG-GI recapitulated known genetic interactions and yielded new condition-dependent genetic interactions. We validated and further explored a subnetwork of condition-dependent genetic interactions involving , and genes encoding the Shu complex, and inferred that loss of the Shu complex leads to an increase in the activation of the checkpoint protein kinase Rad53.
条件依赖性遗传相互作用可以揭示在标准培养条件下不明显的基因之间的功能关系。依赖于机器人操纵双突变株阵列的最先进的酵母遗传相互作用作图技术，不容易扩展到多条件研究。在这里，我们描述了用于绘制遗传相互作用的条形码融合遗传学（BFG-GI），通过“派对”交配产生的双突变株也可以用于生长监测以检测遗传相互作用。通过使用位点特异性重组将两个 DNA 条形码融合在一起，每个条形码代表一个特定的基因缺失，BFG-GI 通过下一代测序实现了双突变体的多路定量跟踪。我们将 BFG-GI 应用于包括甲基甲磺酸（MMS）、4-硝基喹啉 1-氧化物（4NQO）、博来霉素、zeocin 以及其他三种 DNA 损伤环境在内的 9 种不同条件下的 DNA 修复基因矩阵中。BFG-GI 再现了已知的遗传相互作用，并产生了新的条件依赖性遗传相互作用。我们验证并进一步探索了涉及 、 和编码 Shu 复合物的基因的条件依赖性遗传相互作用的子网，推断 Shu 复合物的缺失导致检查点蛋白激酶 Rad53 的激活增加。
