Journal of Data Mining in Genomics & Proteomics

Experimental Approaches for Studying Genomic Alterations in Yeast

Qian Zheng^* Department of Biochemistry and Molecular Biology, National Cheng Kung University Hospital, Tainan, Taiwan
^*Correspondence: Qian Zheng, Department of Biochemistry and Molecular Biology, National Cheng Kung University Hospital, Tainan, Taiwan, Email:

Received: 21-Aug-2024, Manuscript No. JDMGP-24-27262; Editor assigned: 23-Aug-2024, Pre QC No. JDMGP-24-27262 (PQ); Reviewed: 06-Sep-2024, QC No. JDMGP-24-27262; Revised: 13-Sep-2024, Manuscript No. JDMGP-24-27262 (R); Published: 20-Sep-2024, DOI: 10.4172/2153-0602.24.15.351

Description

Yeast, particularly Saccharomyces cerevisiae (budding yeast) and Schizosaccharomyces pombe (fission yeast), has been a model organism for decades in molecular biology, genetics and genomics. Its eukaryotic structure, simple genetic makeup, rapid growth and the ease of genetic manipulation make it ideal for studying various biological processes, including genomic alterations. Genomic alterations, such as mutations, chromosomal rearrangements and gene amplifications, are central to understanding evolution, disease progression (e.g., cancer) and industrial biotechnology applications. This article describes the several experimental approaches used to study genomic alterations in yeast, clarify the molecular tools and methodologies that have advanced our understanding of yeast genetics and genome dynamics.

Mutagenesis for inducing genomic alterations

Mutagenesis is one of the fundamental techniques for studying genomic alterations. This process involves introducing random or targeted changes in the yeast genome to analyze their effects on cellular function, genetic stability, or adaptation.

Chemical mutagenesis: Chemicals such as Ethyl Methanesulfonate (EMS) or nitrosoguanidine can induce random mutations by causing base-pair changes, leading to point mutations. These chemicals often result in single-nucleotide alterations, which can help researchers map genes responsible for specific phenotypes by screening for mutants with altered traits.

UV and ionizing radiation: Ultraviolet (UV) light or ionizing radiation is also used to induce mutations, primarily through the creation of pyrimidine dimers or breaks in the DNA strand. This type of mutagenesis can cause deletions, insertions, or chromosomal rearrangements, allowing the study of larger-scale genomic changes.

Transposon mutagenesis: Transposons, which are mobile genetic elements, can be inserted into the yeast genome to disrupt gene function. This technique is particularly useful for generating loss-of-function mutations, facilitating the identification of essential genes and the study of non-essential genomic regions.

CRISPR-Cas9 for precision genome editing

The advent of CRISPR-Cas9 technology has revolutionized genetic manipulation in yeast. It allows researchers to introduce precise genomic alterations, ranging from single-nucleotide changes to large chromosomal modifications.

Gene knockouts: By designing specific guide RNAs (gRNAs) that target a gene of interest, the Cas9 protein can create Double- Strand Breaks (DSBs) at specific locations in the genome. The cell repairs these breaks through Non-Homologous End Joining (NHEJ), leading to insertions or deletions (indels) that disrupt gene function. This approach allows researchers to study the impact of gene loss on cell survival, metabolism, or stress responses.

Homology-Directed Repair (HDR): When provided with a donor DNA template, yeast cells can repair DSBs using HDR, resulting in precise gene modifications. Researchers can introduce specific point mutations, replace entire genes, or add new sequences, enabling detailed studies of gene function, protein domains and regulatory elements.

Multiplex genome editing: CRISPR-Cas9 also allows for the simultaneous editing of multiple genomic loci, making it possible to study complex traits influenced by multiple genes or pathways. In yeast, this has enabled large-scale genome engineering projects and synthetic biology applications.

High-throughput sequencing for detecting genomic alterations

The development of Next-Generation Sequencing (NGS) technologies has transformed the study of yeast genomics, allowing researchers to detect and characterize genomic alterations at an unprecedented scale and resolution.

Whole-Genome Sequencing (WGS): WGS provides a comprehensive view of the yeast genome, enabling the identification of Single-Nucleotide Polymorphisms (SNPs), insertions, deletions and larger structural variations. This approach is especially useful for studying evolutionary dynamics in populations of yeast, identifying spontaneous mutations and exploring the effects of environmental stresses on genomic stability.

RNA Sequencing (RNA-Seq): Although not directly a tool for detecting genomic alterations, RNA-Seq can be used to assess the effects of genomic changes on gene expression. By comparing the transcriptome of wild-type and mutant yeast strains, researchers can identify how specific genomic alterations influence cellular pathways and regulatory networks.

Comparative genomics: By comparing the genomes of different yeast strains or species, researchers can identify conserved or divergent regions, providing insights into how genomic alterations drive evolution. Comparative genomics also helps pinpoint regions of the genome that are prone to instability or rearrangements.

Chromosome engineering and karyotyping

Chromosomal alterations, including rearrangements, duplications and deletions, are significant in yeast biology and can be engineered for experimental purposes.

Tetrad analysis: Yeast undergoes meiosis, producing four spores, which can be analyzed in tetrads. By dissecting tetrads and analyzing the inheritance patterns of genomic alterations, researchers can study chromosomal segregation, gene linkage and recombination rates. This classic approach remains potential for mapping large genomic alterations.

Fluorescence In Situ Hybridization (FISH): FISH is used to visualize specific regions of the yeast genome within intact cells.

By labeling chromosomal loci with fluorescent probes, researchers can observe chromosomal rearrangements, aneuploidy, or translocations, providing insights into genomic instability and chromosomal dynamics.

Pulse Field Gel Electrophoresis (PFGE): PFGE allows for the separation of large chromosomal fragments, enabling the study of gross chromosomal rearrangements or the identification of new chromosomal structures formed through experimental manipulation or spontaneous alteration.

Synthetic lethality and genetic interaction screens

Genetic interaction screens are a potential tool for studying genomic alterations in yeast. Synthetic lethality occurs when mutations in two genes result in cell death, whereas mutations in either gene alone are viable. This approach allows researchers to uncover genetic interactions and identify pathways that buffer genomic alterations.

Synthetic Genetic Array (SGA) analysis: SGA involves systematic crossbreeding of mutant yeast strains to generate double mutants, which are then screened for synthetic lethality. This technique can help identify pathways that compensate for genomic alterations, revealing functional relationships between genes.

Genome-wide deletion libraries: Yeast deletion libraries, in which each gene is systematically knocked out, are another valuable tool for studying the effects of genomic alterations. These libraries allow researchers to assess the fitness of yeast strains with specific deletions, providing insights into gene function and the role of genomic integrity in maintaining cellular health.