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New research paves the way for rapid and scalable genome engineering in yeast

May 29, 2023
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Yellow yeast cells Yellow yeast cells

A research team led by Aashiq H. Kachroo, an assistant professor in the Department of Biology at Concordia University, has developed a ground-breaking method for advanced genetic engineering.

Rapid, Scalable, and Combinatorial Genome Engineering by Markerless Enrichment and Recombination of Genetically Engineered Loci (MERGE) in Yeast, published in Cell Reports Methods, introduces a new technology that enables highly efficient and scalable genome engineering in yeast. Genetic engineering is crucial in various scientific disciplines, including pharmaceuticals, agriculture, and biotechnology.

Mudabir Abdullah  poses in the lab. He is leaning against a table, wearing blue jeans and a white button-down shirt Mudabir Abdullah

No need for selection markers to edit yeast genome

The team, with PhD student Mudabir Abdullah as a lead author and MSc student Brittany Greco as a contributing co-author, tackles the limitations of traditional genetic engineering methods. These methods have been time-consuming and reliant on selectable markers in yeast.

"Only one in 1000 yeast cells will have edited the DNA using current technology. It is like finding a needle in the haystack,” says Abdullah.

Therefore, to identify the cells with edited DNA, selection markers are linked to the edited sites, thus ensuring the identification of these cells. But this process is not scalable and often tedious - a major bottleneck in DNA editing.

To overcome these challenges, the researchers used Cas9, a genetic scissor, snipping DNA at specific locations as an alternate for selection markers.

“CRISPR-Cas9 system generates targeted and often lethal breaks in yeast’s DNA. However, if the DNA is modified such that the CRISPR-Cas9 no longer recognizes the sequence, those yeast cells can live. Therefore, yeast cells with unmodified DNA are killed, similar to the cells without antibiotic resistance markers. It was fascinating to see the experiment work at near 100 percent precision,” says Abdullah.

Brittany Greco is wearing a black sweater over a wite shirt. She has long brown hair. Brittany Greco

A highly efficient gene drive

By a clever merger of CRISPR-Cas9 and yeast genetics, the team developed a powerful genome engineering platform. Yeast reproduces by two cells mixing both DNAs, Cas9 snips at the unedited locations, triggering yeast's efficient recombination to repair the DNA break with an edited DNA sequence resistant to further cutting by Cas9. This serves as a gene drive that efficiently eliminates specific DNA sequences from the yeast's genome.

Gene drive is a technique that alters the inheritance of specific traits within a population. “It acts like a 'fast lane' that rapidly spreads through the population, almost like an infectious trait,” Kachroo explains. The breakthrough technology consolidates several individual DNA edits into the yeast genome with unprecedented speed and scalability, providing a platform for large-scale genetic manipulation at Concordia’s Genome Foundry.

The implications of this breakthrough are vast. “By streamlining the genome engineering process in yeast, scientists can easily study genetic interactions, build complex biological pathways, and introduce human genetic processes into yeast, as demonstrated by engineering yeast cells to produce yellow carotenoid pigment and the components of the human proteasome,” says Greco.

Learn more about Concordia's Genome Foundry.



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