Where to cut?

Adenine in the "fourth position" increases the accuracy of the genome editing system

Maria Perepechaeva, "First-hand Science"

In recent years, a real stir has been raised around the topic of genome editing, primarily CRISPR/Cas technology – it seems that it is really very simple and accessible. In fact, the pace of use of this technology is much faster than the rate of penetration into the essence of its mechanisms, which is reflected in the low efficiency and accuracy of the method. Scandals and disputes about the permissibility of applying this approach to a person also originate from here. Recently, scientists from the Francis Crick Institute (UK) We have established one of the patterns that will increase the predictability of the results of genomic editing.

The main active element of the currently used CRISPR/Cas genomic editing system is a complex of the so–called guide RNA with the Cas9 protein, which was "borrowed" from a bacterium - pyogenic streptococcus.

This RNA is a synthetic nucleic acid molecule that, like a key to a lock, fits to the part of DNA that needs to be edited. RNA recognizes this fragment and binds to it, after which Cas9 breaks the DNA strand at a place three nucleotides (genetic "letters") away from the end of the gene target. After that, the repair repair system of the cell begins to work at the site of the rupture, and during this process it is possible to purposefully replace a section of the gene if an artificially synthesized suitable donor DNA molecule is delivered to the cell.

Unfortunately, in practice, the guide RNA does not always accurately identify the desired section of DNA, as a result of which the break may occur in a completely different place from where it was planned. In addition, during the "repair" process, potentially dangerous mutations may occur as a result of accidental insertions or loss of nucleotides.

British researchers analyzed the results of many experiments using the CRISPR system aimed at editing about 1.5 thousand targets in 450 genes of the human genome. It turned out that the result of editing largely depends on the nucleotides located directly near the incision site. The nucleotide, the fourth from the break point, is especially important: if it is adenine, the editing accuracy will be high, in the case of guanine, the risk of undesirable rearrangements will be high.

Thus, the very selection of the cut location of the target DNA fragment, taking into account its nucleotide environment, will make genome editing much more predictable, and therefore safer and more promising.

Article by Chakrabartiet al. Target-Specific Precision of CRISPR-Mediated Genome Editing is published in the journal Molecular Cell.

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