Induction of new allelic variant of AhFAD2B gene in peanut cultivar, GG20 through CRISPR/Cas9-mediated mutagenesis

Abstract

The ratio of oleic acid to linoleic acid (O/L) is crucial for determining the shelf life of peanut oil. In peanuts, the expression of the Fatty Acid Desaturase 2 (FAD2) enzyme, which converts oleic acid to linoleic acid, is regulated by two homeologous genes called AhFAD2A and AhFAD2B. It has been observed that the Indian peanut cultivar GG20 does not possess natural mutations in the AhFAD2B gene. We successfully introduced a construct called CRISPR_GG20_AhFAD2B into the de-embryonated cotyledon of the GG20 plant. The purpose of this construct was to utilize the CRISPR-Cas9 gene editing technology to modify the AhFAD2B gene. Genotyping analysis of the plants that were potentially transformed with the construct confirmed that the target-specific editing had occurred in the AhFAD2B gene. Subsequently, the edited GG20 plants were phenotypically evaluated to assess the fatty acid composition in the peanut kernels. The results showed a notable increase in the O/L ratio, which rose from 3.1 in the control to 7.3 in the edited GG20 plants. This implies that the gene editing technique successfully enhanced the oleic acid to linoleic acid ratio in the peanuts, potentially improving the shelf life of the resulting peanut oil.