This finding could increase the number of situations in which CRISPR/Cas9 can be used. Different Cas9 enzymes recognise specific DNA sequences (known as PAM sequences) and can only modify the sites near this sequence. This limitation means that the most commonly used type of Cas9 can only target around one in 16 sites in the genome.
'That's been a real limitation,' said Professor David Liu of Harvard University, Massachusetts, who led the research published in Nature last week. 'If you're trying a CRISPR application that requires very precise positioning of [Cas9] you need to be lucky to have a PAM in just the right place.'
The research team used a method first developed in Professor Liu's lab in 2011 to rapidly evolve Cas9, creating a new variant – xCas9 – that can target around a quarter of the sites in the genome.
'There are numerous efforts to develop CRISPR technologies for clinical and research applications, and we hope that this work increases the likelihood that a particular site of interest is accessible,' explained Professor Liu.
Although they were intending to only broaden the number of PAM sequences that could be targeted, the team found that xCas9 is more precise than other types of the enzyme: an unexpected result, as it was thought that an enzyme that could target more PAM sites would also be less accurate.
'Scientifically, the increased specificity was the biggest shock,' said Professor Liu. It isn't clear yet why xCas9 is more precise. 'If you ask me for a detailed mechanistic explanation for why that is, my answer is, "I don't know".'
'I am amazed that the new Cas9 has both broader PAM recognition and higher specificity,' said Dr Stanley Qi, from Stanford University, California, who was not involved in the study. 'That's amazing biology.'
The full potential of xCas9 as a genome-editing tool will only be clear once it has been tested further: the team tested it on only a few dozen sites in the genome, compared to thousands that the standard Cas9 has been shown to target.
'The hope is that, eventually, we'll have a big bookshelf of Cas9 or other CRISPR enzyme solutions,' says Professor Liu. 'Our efforts in this line of research are never really over until we can access every possible site in the human genome.'