Regeneration and Agrobacterium-mediated genetic transformation of twelve <i>Eucalyptus</i> species

Abstract

<italic>Eucalyptus</italic> is a genus of over 900 species and hybrids, and many of them are valuable fast-growing hardwoods in the world. Due to its economic importance, <italic>Eucalyptus</italic> is one of the early tree species whose genomes were deciphered. However, the lack of efficient genetic transformation system severely restricts the functional genomic research on the plant. The success of <italic>Eucalyptus</italic> regeneration and transformation depends greatly on the genotypes and explants. In this study, we systematically screened 26 genotypes from 12 <italic>Eucalyptus</italic> species with an attempt to obtain <italic>Eucalyptus</italic> genotypes with high regeneration potential. We developed two common regeneration media that can be applied to most tested <italic>Eucalyptus</italic> genotypes for both seeding hypocotyls and cloned internodes as explants. Then, we implemented DsRed2 as a visual marker for genetic transformation efficiency test. Our results suggest that <italic>E. camaldulen</italic> and <italic>E. robusta</italic> are amenable for genetic transformation. Finally, we successfully set up a stable <italic>Agrobacterium</italic>-mediated genetic transformation procedure for both <italic>E. camaldulen</italic> and <italic>E. robusta</italic> using seeding hypocotyls and cloned internodes respectively. Taken together, our study provides valuable means for vegetative propagation, gene transformation, CRISPR based gene mutagenesis, activation and suppression, as well as functional characterization of genes in <italic>Eucalyptus</italic>.