Geminivirus-Mediated Genome Editing in Potato Using Sequence-Specific Nucleases

Genome editing using sequence-specific nucleases (SSNs), such as zinc finger nucleases (ZFNs), TALENs, and CRISPR/Cas is rapidly being improved for genetic engineering in crops. While non-homologous end-joining (NHEJ) has been the preferred DNA repair in plants, gene targeting via homologous recombination (HR) is more precise and allows incorporation of new sequence, and could be a powerful tool for directed DNA repair.

The team of Nathaniel M. Butler from Michigan State University used a geminivirus replicon (GVR) to deliver SSNs targeting the potato ACETOLACTATE SYNTHASE1 (ALS1) gene and repair templates designed to add herbicide-inhibiting point mutations within the ALS1 locus. The team used a geminivirus replicon due to its ability to produce a DNA replicon which can act as a repair template, as well as its relatively large carrying capacity.

Transformed events modified with GVRs held point mutations capable of supporting a reduced herbicide susceptibility phenotype, while plants modified with conventional T-DNAs were just similar to wild-types. Regeneration of transformed events improved detection of point mutations that supported a stronger reduced herbicide susceptibility phenotype.

These results show the effective use of geminiviruses in genome editing, and describes a new approach to gene targeting in vegetatively propagated species.

For more on this study, read the article in Frontiers in Plant Science.