Synthetic Sc2.0 yeast strains contain hundreds to thousands of loxPsym recombination sites that allow restructuring of the Saccharomyces cerevisiae genome by SCRaMbLE. Thus, a highly diverse yeast population can arise from a single genotype. The selection of genetically diverse candidates with rearranged synthetic chromosomes for downstream analysis requires an efficient and straightforward workflow. Here we present loxTags, a set of qPCR primers for genotyping across loxPsym sites to detect not only deletions but also inversions and translocations after SCRaMbLE. To cope with the large number of amplicons, we generated qTagGer, a qPCR genotyping primer prediction tool. Using loxTag-based genotyping and long-read sequencing, we show that light-inducible Cre recombinase L-SCRaMbLE can efficiently generate diverse recombination events when applied to Sc2.0 strains containing a linear or a circular version of synthetic chromosome III.

中文翻译：

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用于识别高度混乱的合成酵母的优化基因分型工作流程

合成的 Sc2.0 酵母菌株含有数百至数千个loxPsym重组位点，允许通过 SCRaMbLE重组酿酒酵母基因组。因此，高度多样化的酵母群体可以由单一基因型产生。选择具有重新排列的合成染色体的遗传多样性候选物进行下游分析需要高效且简单的工作流程。在这里，我们介绍了 loxTags，这是一组 qPCR 引物，用于跨loxPsym位点进行基因分型，不仅可以检测缺失，还可以检测 SCRaMbLE 后的倒位和易位。为了应对大量的扩增子，我们生成了 qTagGer，一种 qPCR 基因分型引物预测工具。使用基于loxTag的基因分型和长读长测序，我们证明光诱导Cre重组酶L-SCRaMbLE在应用于含有线性或环状合成染色体III的Sc2.0菌株时可以有效地产生多种重组事件。