Researchers at Harvard’s Wyss Institute for Biologically Inspired Engineering have created a new gene-editing tool that can allow scientists to perform millions of genetic experiments simultaneously. They call it the Recombinant Library of Retron (RLR) technique, and it uses segments of bacterial DNA called retons that can produce single stranded DNA fragments.
When it comes to gene editing, CRISPR-Cas9 is probably the best known technique today. It’s been making waves in the scientific world over the past few years, giving researchers the tool they need to be able to easily edit DNA sequences. It is more precise than the techniques previously used, and it has a wide variety of potential applications, including vital treatments for various diseases.
However, the tool has major limitations. It might be difficult to deliver CRISPR-Cas9 materials in large numbers, which remains a problem for studies and experiments, for example. In addition, the operation of the technique can be toxic to cells, as the Cas9 enzyme – the molecular “scissors” responsible for cutting DNA strands – often cuts non-target sites as well.
CRISPR-Cas9 physically cuts DNA to incorporate the mutant sequence into its genome during the repair process. During this time, the retons can introduce the mutant DNA strand into a replicating cell, so that the strand can be incorporated into the DNA of the daughter cells. In addition, the reton sequences can serve as “bar codes” or “name tags”, allowing scientists to track individuals in a pool of bacteria. This means that they can be used for genome editing without damaging native DNA, and they can be used to perform multiple experiments in one large mix.
Scientists at the Wyss Institute tested RLR on E. coli bacteria and found that 90 percent of the population incorporated the retron sequence after making some adjustments. They were also able to prove how useful it can be in massive genetic experiments. During their tests, they were able to find antibiotic resistance mutations in E. coli by sequencing the barcodes of the retons instead of sequencing individual mutants, which makes the process much faster.
the study co-first author Max Schubert, explained:
“RLR allowed us to do something impossible to do with CRISPR: we randomly cut out a bacterial genome, turned those genetic fragments into single-stranded DNA in situ, and used them to screen millions of sequences simultaneously. RLR is a simpler, more flexible gene editing tool that can be used for highly multiplexed experiments, which eliminates the toxicity often seen with CRISPR and improves researchers’ ability to explore genome-level mutations …
For a long time, CRISPR was just seen as the odd thing bacteria did, and figuring out how to harness it for genome engineering changed the world. Retrons are another bacterial innovation that could provide important breakthroughs as well. “
There is still work to be done before RLR can be widely used, including improving and standardizing its edit rate. The team believes, however, that this can “lead to new, exciting and unexpected innovations”.
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