Yeast genetic interaction screens in the age of CRISPR/Cas.
Yeast genetic interaction screens in the age of CRISPR/Cas.
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DOI：
10.1007/s00294-018-0887-8
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发表时间：
2019-04
期刊：
影响因子：
2.5
通讯作者：
Peccoud J
中科院分区：
文献类型：
--
作者：
Adames NR;Gallegos JE;Peccoud J
酿酒酵母易于进行正向和反向遗传学，沿着其稳定的单倍体状态和较短的世代时间，使这种芽殖酵母成为遗传学的完美真核细胞模型。使用芽殖酵母进行反向遗传学的主要优点是这种生物体的高效同源定向修复，允许简单地通过引入与染色体靶同源的DNA进行精确的基因组编辑。虽然基于质粒和PCR的基因组编辑工具非常有效，但它们依赖于靶序列附近罕见的自发DNA断裂。因此，它们一次只能生成一个基因组编辑，并且该编辑必须与选择标记相关联。然而，CRISPR/Cas技术足够有效，可以在一个步骤中实现无标记和多重编辑。这些特征使得CRISPR/Cas在合成生物学和代谢工程应用中的酵母菌株工程中很受欢迎，但它尚未广泛用于遗传筛选。在这篇综述中，我们严格审查了在系统遗传相互作用筛选中产生多突变菌株的不同方法，并讨论了CRISPR/Cas补充或改进这些方法的潜力。
The ease of performing both forward and reverse genetics in Saccharomyces cerevisiae, along with its stable haploid state and short generation times, has made this budding yeast the consummate model eukaryote for genetics. The major advantage of using budding yeast for reverse genetics is this organism’s highly efficient homology-directed repair, allowing for precise genome editing simply by introducing DNA with homology to the chromosomal target. Although plasmid- and PCR-based genome editing tools are quite efficient, they depend on rare spontaneous DNA breaks near the target sequence. Consequently, they can generate only one genomic edit at a time, and the edit must be associated with a selectable marker. However, CRISPR/Cas technology is efficient enough to permit markerless and multiplexed edits in a single step. These features have made CRISPR/Cas popular for yeast strain engineering in synthetic biology and metabolic engineering applications, but it has not been widely employed for genetic screens. In this review, we critically examine different methods to generate multi-mutant strains in systematic genetic interaction screens and discuss the potential of CRISPR/Cas to supplement or improve on these methods.
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