Without cuts
New gene editing technology does not cut DNA
Georgy Golovanov, Hi-tech+
CRISPR, a gene editing tool, has revolutionized biology, but it is not without its drawbacks. Harvard scientists have demonstrated an alternative system of Retron Library Recombining (RLR), which does not cut sections of DNA and is able to process millions of mutations and bacterial cells simultaneously. It will be possible to use it in cases when CRISPR is not suitable for one reason or another.
Article by Schubert et al. High-throughput functional variant screens via in vivo production of single-stranded DNA is published in the journal PNAS – VM.
The CRISPR tool acts as a genetic scissors, cutting off and replacing sections of the genome of living cells. The system finds a specific DNA sequence, then uses the Cas9 enzyme to cut it in the right place. And the cell that carries out the repair work receives instructions to use a different sequence instead of the original one. This is how editing happens.
However, in some cases, DNA cutting is fraught with problems – side effects, errors and the inability to make a large number of corrections at a time. In addition, it can be difficult to track what effects these mutations have. The new Retron Library Recombining tool is trying to fill in these gaps, according to a press release from the Wyss Institute Move over CRISPR, the retrons are coming. Its main difference is that it does not cut DNA at all. Instead, the RLR inserts a new segment while the cell reproduces its genome before dividing.
This is done with the help of retrons, segments of bacterial DNA that produce elements of single-stranded DNA (ssDNA). Scientists have used the self-defense mechanism that bacteria use to check if they have been infected by a virus.
"We realized that the retrons should give us the opportunity to produce ssDNA inside the cells that we want to edit, instead of introducing them into the cell from the outside, and without harm to DNA," explained Daniel Goodman, one of the co–authors of the article published in the journal PNAS.
Other advantages of RLR technology include its easy scalability – millions of changes can be made at a time, and the proportion of edited cells increases over time, as cells replicate. In addition, the sequence of retrons can be tracked as a barcode. This gives biologists the ability to easily monitor edited cells.
The system was tested on E. coli, which was injected with antibiotic resistance genes. After just 20 generations, over 90% of the population of these bacteria inherited the desired DNA sequence. And thanks to the barcode, scientists were able to track changes in genes.
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