RALEIGH- In a new study, researchers at North Carolina State University characterize a range of molecular tools for rewriting – not just editing – large chunks of an organism’s DNA, based on CRISPR-Cas systems paired with “auto Selfish genetic “stoppers” called transposons.
Researchers are studying various type IF CRISPR-Cas systems and designing them to add genetic cargo – up to 10,000 additional letters of genetic code – to the transposon cargo to make desired changes to a bacterium – in this case, E.coli.
The results expand the CRISPR toolbox and could have significant implications in manipulating bacteria and other organisms at a time when the need for flexible genome editing in therapeutics, biotechnology and more sustainable and efficient agriculture is necessary.
Bacteria use CRISPR-Cas as an adaptive immune system to resist attacks from enemies like viruses. These systems have been adapted by scientists to delete or cut and replace specific genetic code sequences in a variety of organisms. The new discovery shows that exponentially greater amounts of genetic code can be moved or added, potentially increasing the functionality of CRISPR.
“In nature, transposons have co-opted CRISPR systems to selfishly move through an organism’s genome to help itself survive. We in turn co-opt what happens in nature by embedding transposons with a programmable CRISPR-Cas system that can move the genetic cargo we design to perform certain functions,” said Rodolphe Barrangou, Todd R. Klaenhammer Emeritus Professor of Food. . , Bioprocessing and Nutrition Sciences at NC State and corresponding author of a paper describing the research.
“Using this method, we have shown that we can design genomes by moving pieces of DNA up to 10,000 letters long,” Barrangou said. “Nature already does this – bioinformatics data shows examples of up to 100,000 genetic letters being moved by transposon-based CRISPR systems – but now we can control and engineer it using this system.
Breakthrough in gene editing: NC State researchers use CRISPR to turn the tide on bacteria
“To complete the hitchhiking analogy, we design the hitchhiker to bring certain luggage or goods into the car to deliver some type of payload when the car arrives at its destination.”
The study shows that the researchers prove the effectiveness of the method both in vitro on the laboratory bench and in vivo in E.coli. The researchers selected 10 different transposons associated with CRISPR to test the effectiveness of the method. The approach worked with all 10 transposons, although their efficiency varied depending on factors such as temperature and the size of the transposon’s loading charge.
“It was exciting to see that all of the systems we tested were functional after reconstructing them into genome editing tools from their native biological forms,” said Avery Roberts, NC State graduate student and first author. of the study. “We have discovered new features of these systems, but there are likely many more relevant discoveries and applications to come as the field evolves at a rapid pace.”
The research also showed that the method could be used with different transposons at the same time.
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“Instead of a single gene – as is the case with other CRISPR systems like the more familiar Cas-9 type II system – we can bring a whole metabolic pathway to incorporate a whole new set of functions into a organism,” Barrangou said. “In the future, this could mean providing plants with more flexible resistance to disease or drought, for example.”
“We are excited about these findings and see the potential for applying these newly discovered systems in crop plants to accelerate the development of more resistant and higher yielding varieties,” said Gusui Wu, Global Head of Plants Research. seeds for Syngenta Seeds.
Barrangou and Wu add that the work in this study provides an excellent example of public-private partnerships that are driving scientific discovery and training the workforce of tomorrow.
The paper appears in Nucleic acid research. Funding was provided by Syngenta Seeds. Co-authors of the paper include NC State graduate student Avery Roberts and former NC State Ph.D. student Matthew Nethery.
Note: This story was originally published on this site: https://news.ncsu.edu/2022/11/genetic-hitchhikers-directed-using-crispr/
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