Chalcone synthase (CHS), coded by the elm gene super-family, is a key enzyme in flavonoid biosynthesis. Two independent promoters was isolated for chsA, named PchsA-L (550 bp) and PchsA-S (354 bp) (GenBank accession number EF199747 and EF199748 respectively), from the genomic DNA of Petunia hybrida. PchsA-L differs with PchsA-S mainly in that PchsA-L has a 182 bp fragment from 38 similar to 269 bp, and the sequence 103 similar to 201 bp has the characteristics of a typical intron. Both promoter sequences contain conserved sequences of TATA box, CCAAT box, cap site (CCATAA), and the flower-specific promoter sequences TACPyAT box, anther box (TAGAAGTGACAGAAAT), G-box (CACGTG), box 1 element (ATGTCACGTGCCATC) and box2 element (TGTGTTGAAGGTTTGCTA). The petunia plant used for promoter cloning was a diploid with 14 chromosomes. Southern blotting showed that both promoters had multiple copies in the genome. The two promoters segregated in the offspring but the segregation did not meet the ratio of 1 : 2 : 1. qRT-PCR analysis showed no significant difference in chsA gene expression in non-UV-treated and UV-treated floral organ of plants with one or both promoters. The expression of chsA in the UV-treated seedling leaves was increased compared with UV-treated floral organ, and PchsA-L-driven chsA expression in the UV-treated seedling leaves was very significantly increased than that driven by PchsA-S while there was no chsA gene expression in non-UV-treated seedling leaves. The results show the presence of two independent promoters PchsA-L and PchsA-S for chsA in the petunia genome; the 182 bp intron-like sequence in PchsA-L promoter could significantly increased chsA gene expression in UV-treated seedling leaves.