Abstract: Objective: Phloridzin 2-O-glucosyltransferase (P2’-GT) is the key enzyme involved in the final step in the phloridzin biosynthetic pathway, catalyzing transformation of phloretin into phlorizin. The purpose of this study was to clone a P2’-GT gene from Fuji apple and to investigate bioinformatics analysis of the protein encoded by the gene and its expression pattern. Methods: RNA was extracted from apple peel and was used to synthesize a cDNA template by reverse transcription. Specific primers were designed based on the P2’-GT reporter gene for PCR amplification and sequencing. Bioinformatic analyses were performed using online software (Pro Param tool and TMHMM). The differential transcript levels of P2’-GT in different organs of apple varieties at different growth stages were detected using real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR). Results: The cDNA was 1 452 bp long, encoding 483 amino acid residues. The encoded protein was unstable with a molecular mass of 53.6 kDa and a theoretical PI of 5.76. In addition, the protein had no obvious transmembrane structure. Its secondary structure consisted of α-helix, random coil and extended strand. The protein tertiary structure exhibited high similarity to hydroquinone glucosyltransferase (41.32%). Phylogenetic analysis indicated that P2’-GT had the highest homology to Pyrus bretschneideri glucosyltransferase. The expression level of this gene was high in apple peel but extremely low in leaves and roots and zero in apple pulp. P2’-GT gene expression was related to the growth period; transcript level was rarely detectable in the early stage, but increased rapidly to a maximum in the middle stage and then decreased by 50% in the maturation stage. Significant differences in the transcript level of this gene were found among three different varieties, with the highest and lowest level being observed in Granny Smith and Gala, respectively. This research provides theoretical support for the role of P2’-GT genes in regulating the synthesis of phloridzin.

Key words: phloridzin, phloridzin 2’-O-glycosyltransferase, cloning, bioinformatics, real-time fluorescence quantitative polymerase chain reaction (RTFQ-PCR)