Applications of Genome Editing in Yeast with an Example of Tup1 Mutants Construction

Abstract

Genome editing, which allows for the artificial modification of genes in organisms, has received significant attention in recent years. Currently, various genome editing tools have been developed based on different principles. ZFNs (zinc finger nucleases), TALENs (transcription activator-like effector nucleases), and CRISPR-Cas9 are the three genome editing technologies that are predominantly utilized. This article reviews the principles and characteristics of these three genome editing tools and summarizes the advantages and disadvantages of each of them accordingly. Genome editing is also playing an increasingly important role in yeast, opening up unforeseen applications for human beings. Tup1, a crucial transcriptional corepressor factor in yeast, is associated with multiple metabolic pathways and plays a key role in gene expression regulation. Currently, our understanding of how Tup1 regulates gene transcription remains limited. To gain a better understanding of the detailed mechanisms of Tup1, genome editing can be utilized to deepen our knowledge of its function. Therefore, this article proposes a method that utilizes CRISPR-Cas9 to construct mutant variants of Tup1 in yeast. The main process of this method involves introducing plasmids containing artificially designed sgRNA and Cas9 protein sequences into yeast cells, allowing them to express and edit the yeast Tup1 sequence, ultimately generating yeast Tup1 mutants. This method allows for the efficient construction of engineered Tup1 mutants, facilitating further research on Tup1.