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Key Message: PbMYB1L enhances the cold tolerance and anthocyanin accumulation of transgenic Arabidopsis by regulating the expression of genes related to the cold-responsive genes pathway and anthocyanin synthesis pathway. MYB transcription factors (TFs) have been demonstrated to play diverse roles in plant growth and development. In the present study, we identified a novel R2R3-MYB transcription factor, PbMYB1L, from the peel of 'Red Zaosu' pear (Pyrus bretschneideri), which was induced by cold stress and acted as a positive regulator in anthocyanin biosynthesis. Notably, the transgenic Arabidopsis lines exhibited enhanced tolerance to cold stress. Compared to the Arabidopsis wild-type plants, the transgenic lines displayed longer primary roots and reduced reactive oxygen species (ROS) levels including O2−, hydrogen peroxide (H2O2), and malondialdehyde (MDA). Furthermore, significant upregulation of key cold-responsive genes AtCBF1, AtCBF2, AtCBF3, AtCBF4, and AtKIN1 was observed in the transgenic plants under cold stress conditions compared to wild type. Arabidopsis plants overexpressing PbMYB1L had significant anthocyanin accumulation in leaves after cold treatment with quantitative results indicating higher expression of anthocyanin structural genes compared to wild type. These findings suggest that PbMYB1L not only plays a vital role in conferring cold tolerance but also acts as a crucial regulator of anthocyanin biosynthesis. [ABSTRACT FROM AUTHOR]