To quickly and inexpensively detect wheat transformant plants that contain the mutation in our favorite gene generated by CRISPR, in NGPlab, we have identified restriction enzyme sites lost due to the mutation. So, if the CRISPR-Cas9 machinery managed to generate the mutation in our gene of interest, only «1 band» will be seen in an agarose gel, which cannot be digested by the restriction enzymes and will correspond to a mutant plant!!!!. This is opposite to what is observed in wild-type plants, where you will see the presence of «two bands» generated by the enzyme cutting. This way, we verify only the possible mutants by direct sequencing and optimize the lab’s resources.
Plant Nutrition and Genomics Lab 