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Botrytis cinerea is considered to be the second most destructive fungal pathogen worldwide, causing severe pre‐ and postharvest losses in cut roses. However, to date, no systematic research on its characteristics in cut roses has been reported. In our study, a total of 100 isolates from cut roses from Yunnan, China, were analysed. A combination of morphological characteristics and phylogenetic analysis of RPB2 revealed that 100 isolates were of the species B. cinerea. These isolates were pathogenic on unwounded detached rose petals. Pathogenicity was evaluated according to the size of petal spots and categorized into grades 0–5, from weak to strong. The inhibition rate varied greatly following the addition of procymidone (21.9%–100%) and cyprodinil (25.0%–92.3%). Analysis of Bc‐hch sequences revealed all isolates belong to phylogenetic Group II. According to the presence or absence of transposable elements, 87, 5, 4 and 4 isolates were identified as transposa, vacuma, Boty‐only and Flipper‐only types, respectively. Detection of mating type indicated that all isolates were heterothallic with 45% belonging to MAT1‐1 and 55% to MAT1‐2. Isolates were divided into four subpopulations when analysing single‐nucleotide polymorphisms at the genomic level, with a significant difference in pathogenicity between subpopulations 1 and 4. The evolutionary tree indicated that isolates AN‐02, AN‐22 and SM‐C18 were clustered in the root, suggesting an earlier evolutionary time than other isolates; moreover, they all exhibited low pathogenicity (grade 1). This systematic study of the characteristics of B. cinerea will provide significant support for grey mould control and further research. [ABSTRACT FROM AUTHOR]