Researchers have mixed over seven artificial chromosomes that have been made within the lab right into a single yeast cell, leading to a pressure with greater than 50% artificial DNA that survives and replicates equally to wild yeast strains. The staff current the half-synthetic yeast November 8 within the journal Cell as a part of a group of papers throughout Cell, Molecular Cell,and Cell Genomics that showcase the Artificial Yeast Genome Venture (Sc2.0), a worldwide consortium working to develop the primary artificial eukaryote genome from scratch. The staff has now synthesized and debugged all sixteen yeast chromosomes.
“Our motivation is to grasp the primary rules of genome fundamentals by constructing artificial genomes,” says co-author and artificial biologist Patrick Yizhi Cai of the College of Manchester, who can be senior writer of two different papers within the assortment. “The staff has now re-written the working system of the budding yeast, which opens up a brand new period of engineering biology — shifting from tinkering a handful of genes to de novo design and development of total genomes.”
Although bacterial and viral genomes have been synthesized beforehand, this could be the primary artificial eukaryote genome, which introduces the complication of a number of chromosomes. The artificial yeast can be a “designer” genome that differs considerably from the pure Saccharomyces cerevisiae (brewer’s or baker’s yeast) genome on which it’s based mostly.
“We determined that it was necessary to provide one thing that was very closely modified from nature’s design,” says senior writer and Sc2.0 chief Jef Boeke, an artificial biologist at NYU Langone Well being. “Our overarching purpose was to construct a yeast that may educate us new biology.”
To this finish, the researchers eliminated chunks of non-coding DNA and repetitive components that might be thought of “junk,” added new snippets of DNA to assist them extra simply distinguish between synthesized and native genes, and launched a built-in variety generator known as “SCRaMbLE” that shuffles the order of genes inside and between chromosomes.
To extend genome stability, the staff additionally eliminated lots of the genes that encode switch RNA (tRNA) and relocated them to a completely new “neochromosome” consisting solely of tRNA genes. “The tRNA neochromosome is the world’s first utterly de novo artificial chromosome,” says Cai. “Nothing like this exists in nature.”
Because the yeast genome is organized into sixteen chromosomes, the researchers started by assembling every chromosome independently to create sixteen partially artificial yeast strains that every contained 15 pure chromosomes and one artificial chromosome. The subsequent problem was to start combining these artificial chromosomes right into a single yeast cell.
To do that, Boeke’s staff began through the use of a technique paying homage to Mendel’s peas: primarily, the researchers interbred totally different partially artificial yeast strains after which searched amongst their progeny for people carrying each artificial chromosomes. Although efficient, this technique may be very gradual, however the staff step by step consolidated all beforehand synthesized chromosomes — six full chromosomes and one chromosome arm — right into a single cell. The ensuing yeast pressure was greater than 31% artificial, had regular morphology, and confirmed solely slight progress defects in comparison with wild-type yeast.
To extra effectively switch particular chromosomes between yeast strains, the researchers developed a brand new technique known as chromosome substitution that’s mentioned in one other paper within the new assortment. As a proof of idea, they used chromosome substitution to switch a newly synthesized chromosome (chromosome IV, the most important of all of the artificial chromosomes), leading to a yeast cell with 7.5 artificial chromosomes that’s greater than 50% artificial.
When the artificial chromosomes have been consolidated right into a single yeast pressure, the staff detected a number of genetic defects or “bugs” that have been invisible in yeast strains that solely carried one artificial chromosome. “We knew in precept that this may occur — that we’d have an enormous variety of issues that had tiny little results and that, whenever you put all of them collectively, it’d lead to demise by a thousand cuts,” says Boeke.
A few of these bugs have been merely because of the additive impression of getting many tiny defects inside the genome, whereas others concerned genetic interactions between genes on the totally different artificial chromosomes. The researchers have been in a position to map and repair a number of of those bugs and improve the artificial yeast’s health through the use of a technique based mostly on CRISPR/Cas9.
“We have now proven that we will consolidate primarily half of the genome with good health, which means that this isn’t going to be a giant drawback,” says Boeke. “And from debugging, we study new twists on the foundations of life.”
The subsequent step will probably be to combine the remaining artificial chromosomes. “Now we’re simply this removed from the end line of getting all 16 chromosomes in a single cell,” says Boeke. “I prefer to name this the tip of the start, not the start of the tip, as a result of that is once we’re actually going to have the ability to begin shuffling that deck and producing yeast that may do issues that we have by no means seen earlier than.”
This analysis was supported by the Nationwide Science Basis, the Nationwide Institutes of Well being, the Laura and Isaac Perlmutter Most cancers Heart, and Volkswagen Stiftung.
