CRISPR Variants for Gene Editing in Plants: Biosafety Risks and Future Directions

Abstract

The CRISPR genome editing technology is a crucial tool for enabling revolutionary advancements in plant genetic improvement. This review shows the latest developments in CRISPR/Cas9 genome editing system variants, discussing their benefits and limitations for plant improvement. While this technology presents immense opportunities for plant breeding, it also raises serious biosafety concerns that require careful consideration, including potential off-target effects and the unintended transfer of modified genes to other organisms. This paper highlights strategies to mitigate biosafety risks and explores innovative plant gene editing detection methods. Our review investigates the international biosafety guidelines for gene-edited crops, analyzing their broad implications for agricultural and biotechnology research and advancement. We hope to provide illuminating and refined perspectives for industry practitioners and policymakers by evaluating CRISPR genome enhancement in plants.

Keywords: CRISPR variants; biosafety; gene editing; off-target effects; plant genetic improvement.

Figure 1

The critical differences between Cas9 and Cas12a are illustrated here. (A): (a) Cas9 has two endonuclease domains that, when activated, use the RuvC and HNH domains to target the target strand (TS) and non-target strand (NTS). (b) Cas9 needs tracrRNA for the synthesis of mature crRNA. (c) For the cleavage of the target site, the PAM of Cas9 requires NGG-rich areas. (d) Cas9 produces blunt ends while concurrently breaking both the TS and NTS. (B): (a) Cas12a uses RuvC, a single endonuclease domain, to cut the TS and NTS. (b) Cas12a generates mature crRNA without the assistance of tracrRNA. (c) The Cas12a PAM requirements are “TTN/TTTN”, with a preference for AT-rich areas. (d) Cas12a cleaves the NTS first, then the TS, resulting in a double-strand staggered break (sticky ends).