*1.5. Regulatory Framework of CRISPR-Cas-Edited Crops*

During CRISPR-Cas gene editing of plants, Cas cuts the target sequence to produce double-strand breaks, resulting in the loss of gene function. The CRISPR-Cas technique has been utilised to create modifications in the genome that are identical to natural genetic variation [94]. Similarly, in HDR (homologous DNA repair) of CRISPR-Cas-mediated dsDNA fragmentation, exogenously provided homologous DNA sequences are deemed transgenic; however, when the repair template is derived from the genes from the same species and related interbreeding species, the resultant crops are not regarded as transgenic crops [125]. Nevertheless, the regulatory framework regarding NHEJ and HDR-mediated gene editing contains differing definitions.

The United States Department of Agriculture (USDA), FDA, and Environmental Protection Agency state that the removal of transgenic elements in plants by CRISPR-mediated editing is equivalent to crop improvement by conventional breeding programs, and crops generated in this way are thus not considered GMOs for regulatory purposes [126]. In 2016, the USDA approved the marketing of gene-edited waxy corn without exogenous transgenic elements [127], and an *Agaricus bisporus* mushroom with an anti-browning trait obtained from CRISPR-Cas9 editing was exempted from GMO regulatory procedures [128]. This definition of gene-edited crops by the United States regulatory agencies promotes the

genetic improvement of crops and accelerates the introduction of gene-edited crops to the market. Genome editing has emerged as a powerful and elegant technology to develop novel varieties or organisms with desirable traits in tomato, citrus, soybean, sugarcane, camelina, and rice. In May 2020, the USDA–APHIS issued the latest edition of biotechnology regulations that provide three exemptions for genetic modifications in any plant species: (i) resultant changes in DNA after DSB in the absence of an external repair donor template; (ii) single base pair substitution in targeted loci; and (iii) introduction of a known gene that exists in the plant's gene pool.

As a major producer of GM crops, Canada considers gene-edited products such as plants, animal feed, or human food as different from nonedited products, so they must undergo a premarket assessment. In Europe, there are stringent regulations regarding CRISPR/Cas9, and the European Court of Justice has included gene-edited crops in the scope of GM crop regulation [129]. Australia has taken a milder approach, allowing geneediting without the introduction of any foreign genetic material [130]. In Asia, the attitude towards gene-edited crops has eased in China and Japan, and cases of gene-edited crops being planted in the field have been reported [131]. In addition, some countries have regulatory frameworks that are applied case-by-case, considering the breeding methodology used, new traits or characteristics introduced, and evidence of the genetic changes in the final product.
