Using Retrons to Improve Genome Editing Through More Efficient Modification
By: April Carson
The CRISPR genome-editing technology has revolutionized molecular biology over the last decade, allowing researchers to modify genes inside living cells for study or medical purposes. Researchers at the Gladstone Institutes have fine-tuned a second system for more efficient gene editing, employing retrons.
Retrons, according to a study published in Nature Chemical Biology, may be optimized for efficiency and used to modify genes in a wide range of cell types, from fungi to human cells.
“This research really validates retrons as a platform that may be utilized by different species,” says Gladstone Assistant Investigator Seth Shipman, PhD, senior author of the paper. We can now produce more precise changes to genes than before, quickly and efficiently.”
A One-Stop Shop for Gene Modification
The CRISPR-Cas9 system is used in most current gene editing techniques, which entail removing a section of DNA from a cell's genome and replacing it with "template DNA," or new genetic material. The template DNA is introduced as the cell repairs the regions where an old gene has been removed.
The CRISPR template DNA is generally created in a laboratory and then fed into cells from the outside. Cas9, the enzyme that cuts up the genome of a cell, is supplied separately. Neither Cas9 nor the CRISPR template DNA reach every cell, limiting CRISPR gene editing's efficacy.
However, retroviruses do not merely paste themselves into the DNA of their host cells; instead, they act as DNA factories, generating a large number of copies of the template DNA from within cells. Finally, retrons can be included with the rest of the CRISPR components to ensure that cells receive all of the supplies they need for genetic modification at once—the genetic codes for template DNA, Cas9, and chemicals that aid in tracking changes.
“This implies that each cell only has to be introduced to one component,” says Santiago Lopez, a graduate student in the Shipman Lab and lead author of the new study. “This significantly lowers the barrier to experimentation by removing several steps.
Shipman and Lopez developed a retron system that improves gene editing performance.
Retrons are being re-engineered
The CRISPR-Cas9 gene editing system, which is also known as retrons, was discovered in bacteria. Both retrons and CRISPR are bacterial defense mechanisms that modify DNA in response to infections. After the advent of CRISPR gene editing, when scientists co-opted the CRISPR system to target genes in other cell types, some researchers began looking into whether retrons might be used to provide the templates for precise genetic alteration. However, the functions of various sections of the retron's structure and how to modify them to enhance retrons' efficiency with CRISPR have not been clear.
“The retron system was created to help bacteria defend themselves,” adds Shipman, who is also an assistant professor of bioengineering and therapeutic sciences at UCSF. “But we wanted it to do something different – make gene editing templates.”
Shipman's team modified E. coli retrons to generate hundreds of new variants in a recent research. They tested each new variant and discovered a sequence of changes that, together, boosted the amount of template DNA produced by the retron in E. coli cells by 8-to-10 times.
Next, the team tested retrons in S. cerevisiae (Baker's yeast) and human cells cultivated in a lab, and they discovered that the new re-engineered retron system worked in all cases. This was the first proof of retrons' potential use in human tissue and across cell types.
The researchers were able to study how the brain learns through electrical stimulation of the whisker receptors after determining whether or not they are present in the region. This is due to a recent development with their lab’s technique for editing genes in different cell types.
Since the researchers could now precisely control exactly how much template DNA the retrons created, they were also able to demonstrate that when retrons generate a lot of template DNA, gene editing efficacy improves.
“Our research demonstrates for the first time that the more template DNA we can produce, the better genome editing,” says Shipman. “Improved and more accurate gene editing leads to improved genomic medicines and further fundamental study.”
Taking Tools from Bacteria
Retrons, according to Shipman, are suitable for use as a research tool in the lab to modify genes in various cell types. While the platform is not yet ready for human usage, it might also have the ability to help cure genetic abnormalities that lead to disease.
His team will continue to study the potential benefits of retrons for other bacterial strains. Because bacteria come in a variety of shapes and sizes, however, they must figure out how those variations might help them adapt and overcome antibiotic resistance.
“We're taking a broad approach by mining components found in bacteria and domesticating them for our own purposes,” says Shipman. “This has already been quite successful in the area of R&D, but I believe we are only now seeing the full benefits of these technologies in biotechnology.”
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About the Blogger:
April Carson is the daughter of Billy Carson. She received her bachelor's degree in Social Sciences from Jacksonville University, where she was also on the Women's Basketball team. She now has a successful clothing company that specializes in organic baby clothes and other items. Take a look at their most popular fall fashions on bossbabymav.com
To read more of April's blogs, check out her website! She publishes new blogs on a daily basis, including the most helpful mommy advice and baby care tips! Follow on IG @bossbabymav
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