Abstract: [Objective] Anthocyanin is an important pigment in the flower coloration of Prunus mume. R2R3-MYB transcription factor is a key regulator of anthocyanin biosynthesis. In the present study, the PmMYB1 gene has been isolated from P. mume. This gene was analyzed for its sequence characteristics and was transferred into tobacco to verify its biological function. This will lay a foundation for the research on the regulation mechanism of anthocyanin biosynthesis and the molecular breeding of flower color of P. mume.[Method] The PmMYB1 gene was isolated from the petal of P. mume using RT-PCR method. The specific primers for PCR amplification were designed according to the P. mume genome data. Multiple sequence alignment and phylogenetic tree were used to analyze the conserved sequences and predict the biological function of PmMYB1. PmMYB1 gene was transferred into tobacco by constructing expression vector. The changes of phenotype and anthocyanin content in transgenic tobacco were analyzed. In addition, qRT-PCR was used to detect the expression levels of the PmMYB1 and the endogenous anthocyanin biosynthetic genes in different organs of transgenic tobacco. All data were analyzed to identify the function of the PmMYB1 gene.[Result] The CDS sequence of PmMYB1 (729 bp) were obtained by cloning from P. mume. The PmMYB1 protein encoded 242 amino acids. Sequence analysis showed that the protein had a conserved MYB domain and specific motif of anthocyanin-regulating MYB protein, and contained a conserved motif that interacts with the bHLH transcription factor, which might require the co-expression of bHLH protein in the regulation of anthocyanin synthesis. Multiple sequence alignment and phylogenetic analysis revealed that PmMYB1 was highly homologous with other identified anthocyanins-regulating MYB proteins, such as 92% and 91% identify with PcMYB10 from Prunus cerasifera and PaMYB10 from Prunus avium, respectively. Transgenic tobacco assay showed that overexpression of PmMYB1 gene significantly promoted the accumulation of anthocyanin in the corolla of transgenic plants. Compared with the control, the anthocyanin content in the flowers of transgenic plants was higher(P< 0.05), which caused to the deepening of flower color. The qRT-PCR assay showed that the expression level of PmMYB1 gene was higher in the flowers of transgenic plants than that of leaves. The expression level of seven structural genes(NtCHS, NtCHI, NtF3H, NtF3'H, NtDFR, NtANS, and NtUFGT)and two regulatory genes(NtAn1a and NtAn1b)of anthocyanin biosynthesis in the flowers of transgenic lines were obviously up-regulated compared with the control.[Conclusion] Overexpression of PmMYB1 in tobacco significantly promoted anthocyanin accumulation in the flowers and caused to the deepening of flower color of transgenic plants through activating endogenous structural and regulatory genes of anthocyanin biosynthesis to up-regulate their expression in the flowers of transgenic lines. These findings suggest that PmMYB1 plays an important role in the regulation of anthocyanin synthesis.

Key words: Prunus mume, anthocyanin regulated transcription factor, gene cloning, transgenic tobacco, functional identification