What is the available evidence for the range of applications of genome editing as a new tool for plant trait modification and the potential occurrence of associated off-target effects: a systematic map

This is a systematic review that provides an overview of publications in which genome editing was used to change the genome of plants and investigates the occurrence of off-target effects (from January 1996 to May 2018 inclusive). Applications of genome editing techniques (CRISPR/Cas, TALENs, ZNF, meganucleases, ODM and base editors) on agriculturally relevant plants, as well as on model organisms from basic research were documented. Most often, CRISPR/Cas is used to introduce small changes in the plant genomes without providing a repair template (so-called SDN1 applications). A total of 99 different market-oriented applications such as increasing the agronomic value or introducing stress tolerances using genome editing were identified. When investigating off-target effects (OTEs), most of the studies used biased methods that are based on investigating genomic regions that are very similar to the target sequence as predicted by computer programmes (in silico). The gene scissors can cut at these off-target sites with a certain probability causing unintended alterations in the genome.
These off-target sites are then specifically checked after the genome editing procedure whether OTEs have occurred there. In only 252 studies and in 1328 applications of genome editing off-target sites were investigated. Of these, 211 CRISPR/Cas applications searched for OTEs in previously determined DNA areas and 55 OTEs were actually found. Only 9 studies used whole genome sequencing (WGS) procedures to examine the entire genome in an unbiased way. No OTEs that could be associated with the genome editing process were found in these studies. In studies that used TALENs, WGS was carried out in only one study and 3 OTEs were actually found. However, it is unclear whether these were caused by the TALENs or by spontaneously occuring mutations. The authors conclude that, in order to make a clear statement about the occurrence of OTEs when using genome editing techniques, much more unbiased WGS investigations have to be carried out and individual studies regarding the occurrence of OTEs need to be critically analyzed.