Transposon-based CRISPR systems can move up to 100,000 bases

In a new study, researchers at North Carolina State University describe a series of molecular tools to rewrite — not just edit — chunks of an organism's DNA, based on the CRISPR-Cas system, associated with selfish genes known as transposons, "free riders
.
"

The researchers studied different I-F type CRISPR-Cas systems and engineered them to add up to 10,000 additional genetically encoded letters to the vectors of transposons to make the desired changes
to the bacteria, in this case E.
coli.

The discovery expands the CRISPR toolbox that could have a significant impact
on the manipulation of bacteria and other organisms when flexible genome editing is required in therapeutics, biotechnology, and more sustainable and efficient agriculture.

Bacteria use CRISPR-Cas as an adaptive immune system to fend off attacks from enemies such as viruses
.
These systems have been applied by scientists to remove, cut, and replace specific genetic code sequences
in various organisms.
The new findings suggest an exponential increase in the amount of genetic code that can be moved or added, which could increase the functionality of
CRISPR.

"In nature, transposons chose the CRISPR system to selfishly move themselves through the genome of an organism to help themselves survive
.
In turn, by integrating a programmable CRISPR-Cas system with transposons, we can move the genetic cargo that we designed to perform certain functions," said Rodolphe Barrangou, the Todd R.
Klaenhammer Associate Professor of Food, Bioprocessing, and Nutritional Sciences at North Carolina State University and corresponding author
of a paper describing the study.

Barrangou said: "Using this method, we demonstrated that we can engineer genomes
by moving blocks of DNA up to 10,000 letters.
Nature has done just that—bioinformatics data shows that transposon-based CRISPR systems can move up to 100,000 genetic letters—but now we can control and engineer it
by using this system.

"To complete the hitchhiking analogy, we designed hitchhikers to bring specific luggage or cargo into the car, delivering a certain type of payload
when the car arrives at its destination.
"

This study shows that the researchers demonstrated the effectiveness of
this approach both on in vitro benchtops and in E.
coli.
The researchers selected 10 different CRISPR-related transposons to test the effectiveness of
the method.
This method is suitable for all 10 transposons, although their effectiveness varies
depending on factors such as temperature and transposon load size.

Avery Roberts, a graduate student at North Carolina State University and first author of the study, said: "It's exciting that all the systems we tested worked correctly
after reconstructing them from protist morphology into genome editing tools.
We have discovered new features in these systems, but as the field rapidly evolves, more relevant discoveries and applications may emerge.

”

Studies have also shown that this method can be used for different transposons
at the same time.

Barrangou said: "Unlike other CRISPR systems, such as the more familiar type II Cas-9 system, which has only one gene, we can introduce the entire metabolic pathway, integrating a whole new set of functions into the organism
.
In the future, for example, this could mean providing plants with more flexible disease or drought resistance
.
”

Gusui Wu, global head of seed research at Syngenta Seeds Seeds, said: "We are excited about these findings and see the potential to apply these newly discovered systems to crop crops to accelerate the development of more resilient, higher-yielding varieties
.
"

Barrangou and Wu added that the work in this study provides a great example
of public-private partnerships that drive scientific discovery and train the workforce of the future.

Functional characterization of diverse type I-F CRISPR-associated transposons