The Purdue team created a method that uses the protein Argonaute from Natronobacterium gregoryi (NgAgo) and supplied DNA as a guide to enable modification anywhere on the genome, providing new options to potentially improve manufacturing, disease treatment, drug discovery and crop production.
NgAgo is a DNA-guided DNA endonuclease Although previous studies demonstrated that refolded NgAgo does not cut DNA in vitro , consistent with our findings, we establish that soluble NgAgo can, in fact, cleave DNA in vitro. That is, refolded NgAgo may not be fully functional. As we showed that an N-del/D663A/D738A catalytic mutant lacks DNA cleaving activity (Fig. 2e), the catalytic activity is unlikely to be the result of sample contamination. However, we are unable to demonstrate unequivocal guide-dependent cleavage with both double-stranded DNA target and single-stranded DNA target in vitro (data not shown). This may be due to inefficient guide loading, as we observe that N-del co-purifies guides (Fig. 2c).
NgAgo can be repurposed as a DNA editing tool
Our results provide supporting evidence to encourage the development of NgAgo for gene-editing.