Identification and Classification of LEA Family Genes in Orchids and Characterization of Their Role in Callus Formation

The plant late embryogenesis abundant (LEA) proteins are abundant in seeds, play an important role in various abiotic stresses. However, there is still no information on genome-wide identification of LEA genes in orchids and their function in callus formation is almost unknown. In this study, the LEA genes from two orchids (Phalaenopsis equestris and Dendrobium officinale), were genome-wide identified, classified and characterized. A total of 57 and 59 LEA genes were identified in the genomes and these were divided into 8 and 9 groups for P. equestris and D. officinale, respectively. The LEA_1 and LEA_4 genes from P. equestris and D. officinale showed strong expression in seeds, but were significantly down-regulated in flowers and absent in vegetative organs (leaves, stems and roots). In addition, the LEA_1 and LEA_4 genes from D. officinale were abundant in the protocorm-like body (PLB) stage, while weak signals that were detected in in vitro shoots could not be detected in plantlets. The expression of these genes highlights PLBs in orchids are somatic embryos. The DoLEA36 from LEA_4 and DoLEA43 from LEA_1 were further characterized. The GFP signal of the DoLEA36-GFP fusion protein was only detected in the cytoplasm, while the GFP signal of the DoLEA43-GFP fusion protein was detected in both the cytoplasm and nucleus. This indicates that DoEA36 localizes in the cytoplasm while DoLEA43 localizes in both the cytoplasm and nucleus. Both DoLEA36 and DoLEA43 stimulated callus formation in transgenic Arabidopsis. The percentage of callus formation from 35S::DoLEA43 transgenic lines was higher than in wild type plants in two callus induction methods. Our results provide comprehensive information about the LEA gene family in orchids and genetic evidence for the involvement of LEA genes in the induction of callus, which may reveal their positive role in the maintenance of PLBs in orchids.