Cloning whole bacterial genomes in yeast

Area of Science:

• Microbiology
• Molecular Biology
• Genomics

Background:

• Bacterial and archaeal genomes are often comparable in size to DNA molecules successfully cloned in Saccharomyces cerevisiae.
• Yeast (Saccharomyces cerevisiae) provides a robust platform with established tools for DNA manipulation and analysis.

Purpose of the Study:

• To present a strategy for cloning entire bacterial and archaeal genomes as single, circular episomes in yeast.
• To enable the study and manipulation of microbial genomes using yeast-based molecular biology techniques.

Main Methods:

• Cotransformation of yeast spheroplasts with the target genome and a linear yeast vector.
• The yeast vector contains termini homologous to specific regions within the target genome, facilitating integration.
• Selection of successful transformants using auxotrophic markers, followed by screening for clone completeness and size.

Main Results:

• Demonstration of a viable method for the episomal cloning of microbial genomes in Saccharomyces cerevisiae.
• Successful identification and selection of complete genome clones.

Conclusions:

• Episomal cloning of microbial genomes in yeast is feasible and offers a powerful approach for genomic research.
• This technique facilitates subsequent sequencing and detailed analysis of cloned bacterial and archaeal genomes.