Transcriptomic analysis of papaya (Carica papaya L.) shoot explants obtained by leaf- and stem-inoculation methods for adventitious roots induction

Tissue culture has been widely used for the propagation of papaya (Carica papaya L.). Generally, agar-based media supplemented with the appropriate amount of indole-3-butyric acid (IBA) is used for the in-vitro rooting and root elongation of the papaya explants. However, it is usually resulted in a difficulty in adventitious root (AR) formation and turns into callus formation. The AR formation was influenced by miscellaneous factors, however little is known of the related regulative mechanisms. In this study, RNA-seq was performed on samples collected from leaf-inoculation (LI) and stem-inoculation (SI) GZY6 shoot explants to reveal their different rooting phenotypes. Here, we found that the ARs were induced from in GZY6 (79.17%), GZG4 (63.33%), and GZM26 (37.23%) hermaphroditic papaya shoot explants obtained by LI method after 2 weeks of culture in root-induction medium, but calli were formed rather than AR from the explants obtained by SI method in root-induction medium. The SI and LI shoot explant samples of GZY6 genotypes (the rooting percentage was highest in the three genotypes) were used for the transcriptomic analyses. A total of 2498 differentially expressed genes (DEGs) were identified in the LI vs. SI samples and annotated into 73 Gene ontology (GO) terms. The GO analysis revealed that oxidative-stress, defense-response, and stimulus-response GO terms were significantly different between the LI and SI shoot explants and that the genes are involved in anaerobic respiration and reactive oxygen species (ROS) metabolism pathways. Twelve selected anaerobic and ROS metabolic pathways-related DEGs were used to validate the RNA-seq data by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The activity of superoxide dismutase (SOD) and peroxidase (POD) and the concentration of hydrogen peroxide (H2O2) increased in the SI explants. ARs formation and development were significantly higher in the SI explants in perlite medium than in agar medium, when both were not supplemented with IBA. Papaya shoot explants absolutely formed ARs in peat medium after stem bases dipping in 1 mg mL(-1) IBA for 3 s. Our results indicate that the root-induction medium supplemented with agar and IBA would be mostly limited AR emergence and induced callus formation in SI explants, respectively. This study provides basic data for establishing a healthy and functional rooting system for in vitro papaya propagation.