EgENOD93 was first identified in a cDNA microarray study of oil palm tissue culture where it was highly expressed in leaf explants with embryogenic potential. Functional characterization via an RNA interference study of its orthologue in Medicago truncatula demonstrated a significant role of this gene in somatic embryo formation. In this study, EgENOD93 was overexpressed in the important model plant Arabidopsis thaliana to investigate the embryogenic potential of EgENOD93 transgenic Arabidopsis explants compared to explants from control plants (pMDC140 and WT). Experiments using leaf explants revealed higher numbers of regenerated shoots at day 27 in all the homozygous transgenic Arabidopsis cultures (Tg01, Tg02 and Tg03) compared to controls. The expression level of EgENOD93 in Arabidopsis cultures was quantified using reverse transcription quantitative real-time PCR (RT-qPCR). The results supported the overexpression of this gene in transgenic Arabidopsis cultures, with 6 and 10 times higher expression of EgENOD93 in callus at Day 9 and Day 20, respectively. Overall, the results support the role of EgENOD93 in the enhancement of shoot regeneration in transgenic Arabidopsis. This together with the previous results observed in oil palm and Medicago truncatula suggests that ENOD93 plays a key role in the induction of somatic embryogenesis. A similarity to early nodulation-like ontogeny is possible.
KEY MESSAGE: Transcript profiling during the early induction phase of oil palm tissue culture and RNAi studies in a model somatic embryogenesis system showed that EgENOD93 expression is essential for somatic embryogenesis. Micropropagation of oil palm through tissue culture is vital for the generation of superior and uniform elite planting materials. Studies were carried out to identify genes to distinguish between leaf explants with the potential to develop into embryogenic or non-embryogenic callus. Oil palm cDNA microarrays were co-hybridized with cDNA probes of reference tissue, separately with embryo forming (media T527) and non-embryo (media T694) forming leaf explants sampled at Day 7, Day 14 and Day 21. Analysis of the normalized datasets has identified 77, 115 and 127 significantly differentially expressed genes at Day 7, Day 14, and Day 21, respectively. An early nodulin 93 protein gene (ENOD93), was highly expressed at Day 7, Day 14, and Day 21 and in callus (media T527), as assessed by RT-qPCR. Validation of EgENOD93 across tissue culture lines of different genetic background and media composition showed the potential of this gene as an embryogenic marker. In situ RNA hybridization and functional characterization in Medicago truncatula provided additional evidence that ENOD93 is essential for somatic embryogenesis. This study supports the suitability of EgENOD93 as a marker to predict the potential of leaf explants to produce embryogenic callus. Crosstalk among stresses, auxin, and Nod-factor like signalling molecules likely induces the expression of EgENOD93 for embryogenic callus formation.
Acyl-CoA-binding proteins (ACBPs) are involved in binding and trafficking acyl-CoA esters in eukaryotic cells. ACBPs contain a well-conserved acyl-CoA-binding domain. Their various functions have been characterized in the model plant Arabidopsis and, to a lesser extent, in rice. In this study, genome-wide detection and expression analysis of ACBPs were performed on Elaeis guineensis (oil palm), the most important oil crop in the world. Seven E. guineensis ACBPs were identified and classified into four groups according to their deduced amino acid domain organization. Phylogenetic analysis showed conservation of this family with other higher plants. All seven EgACBPs were expressed in most tissues while their differential expression suggests various functions in specific tissues. For example, EgACBP3 had high expression in inflorescences and stalks while EgACBP1 showed strong expression in leaves. Because of the importance of E. guineensis as an oil crop, expression of EgACBPs was specifically examined during fruit development. EgACBP3 showed high expression throughout mesocarp development, while EgACBP1 had enhanced expression during rapid oil synthesis. In endosperm, both EgACBP1 and EgACBP3 exhibited increased expression during seed development. These results provide important information for further investigations on the biological functions of EgACBPs in various tissues and, in particular, their roles in oil synthesis.