| 【Objective】The study is presented to obtain the sequence information of polyphenol oxidase gene family in Volvariella volvacea, and identify their characteristics and expression patterns.【Method】The genes containing tyro sinase domains were found out through transcriptome sequencing, and their primers were designed according to the obtained sequences. By using the tissues at different developmental stages of the v26 strain, as well as the outer membrane,
cap, and stipe of the egg-shaped stage as materials, RNA was extracted and reversely transcribed into cDNA. The cDNAs were used as the template to amplify the polyphenol oxidase gene sequences. The online bioinformatics website was used to predict the physicochemical properties of V. volvacea polyphenol oxidase. The RT-PCR method was used to detect the expression patterns of polyphenol oxidase in different developmental stages and different tissues.【Result】A total of four CDS sequences of PPO genes were cloned from V. volvacea, named as VvPPO1, VvPPO2, VvPPO3 and VvPPO4. The VvPPO1, VvPPO2, VvPPO3 and VvPPO4 CDSs were 1 596 bp, 2 685 bp, 1 593 bp and 2 067 bp in length and encoded 531, 894, 530 and 688 amino acids, respectively. Bioinformatics analysis showed that the proteins encoded by the four genes were hydrophilic proteins. VvPPOs had typical characteristics of PPO protein, holding a tyrosinase central domain which could bind two copper ions. RT-PCR results showed that the 4 genes were expressed in the hyphae, primordium, egg-shaped and mature stages. The expression of VvPPO1, VvPPO3 and VvPPO4 in the fruiting body were higher than that in the hyphae. In the egg-shaped stage, the expression of VvPPO1 and VvPPO4 in the outer membrane were higher than that in the stipe and cap.【Conclusion】A total of four VvPPO genes were obtained in V. volvacea, all of which contained a tyrosinase central domain and two copper-binding sites, and VvPPO genes were expressed in all developmental stages. |
