Rapid production of abundant transgenic pomegranate (Punica granatum) hairy roots

Abstract

Pomegranates (Punica granatum) are known for their high levels of health-beneficial compounds that belong to the hydrolyzable tannin (HT) and flavonoid families. However, a significant gap in our understanding exists of the biosynthetic and regulatory genes related to the accumulation of these compounds. To this end, the induction and transformation of hairy roots presents an exciting opportunity to decipher genetically the functions of candidate genes involved in the HT and flavonoid metabolism. This study aims at improving methods for rapid and abundant hairy root production from pomegranate explants by increasing seed germination rates and expediting the attainment of requisite biomass for multifaceted analyses. We found that treating seeds with sandpaper and concentrated sulfuric acid significantly enhanced their germination rates. Interestingly, no correlation between seed-coat texture and seed germination rate was observed among the 20 pomegranate accessions evaluated in this study. Notably, transferring hairy roots from agar plates to soil promoted rapid root biomass growth compared to maintaining them on agar plates. Root biomass as well as levels of gallic acid (a precursor for HT biosynthesis) and punicalagins (the major HTs in roots) across 15 pomegranate accessions were also analyzed. Three of these accessions exhibited higher seed germination percentages along with augmented biomass and elevated punicalagin levels. These pomegranate accessions emerge as promising germplasm for future genetic transformation and functional genomics studies.