CRISPR-Cas9 come strumento di editing genetico programmabile

The transformation of the CRISPR-Cas9 bacterial immune system into a universal tool for genome editing was a landmark achievement in molecular biology. The key insight was recognizing its potential for being reprogrammed. In its natural form, the Type II system uses three components: the Cas9 protein, a crRNA that contains the targeting sequence, and a tracrRNA that is crucial for crRNA maturation and Cas9 activation. The Doudna and Charpentier labs demonstrated that this system could be simplified. They engineered a single chimeric RNA, which they termed a single-guide RNA (sgRNA), by linking the 3′ end of the crRNA to the 5′ end of the tracrRNA with a synthetic hairpin loop. This sgRNA retained all the necessary functions of the dual-RNA system.

Questa semplificazione è stata rivoluzionaria perché significava che per reindirizzare la nucleasi Cas9 verso un nuovo sito di DNA, era sufficiente sintetizzare un nuovo sgRNA con una diversa sequenza guida di 20 nucleotidi.