Genome editing is widely used across plant species to generate and study the impact of functional mutations in crop improvement. However, transgene integration in plant genomes raises important legislative concerns regarding genetically modified organisms.
Genome editing methods have been developed to introduce precise and predictable genome modifications into plants to achieve the desired features, and they are giving rise to precision breeding techniques that will define the next generation of plant breeding. CRISPR-Cas (clustered regularly interspaced short palindromic repeats) is one of the most sophisticated techniques for engineering crop genomes. Moreover; newly discovered CRISPR-associated tools such as base editors and prime editors have dramatically expanded the scope of genome editing, allowing for the creation of precise nucleotide substitutions and targeted DNA deletions and insertions. Recently, the Cas12b system was also developed for plant genome editing. All of these systems rely upon crRNAs to guide the Cas protein to target sequences.
The general procedure for genome editing in plants can be divided into six steps:
(1) Select the appropriate nuclease based on the target sequence; (2) Construct genome editing vectors; (3) Validate the activity of these vectors using protoplasts (wall-free plant cells released from enzyme-digested tissues; optional step); (4) Deliver genome editing reagents into plant cells; (5) Regenerate genome-edited cells into plantlets via tissue culture; and (6) Screen and genotype the resulting genome-edited plants.
Dr. Md. Monirul Islam
ASRBC, ACI Seed