Cloning and Analysis of Two Promoters of ChalconeSynthase Gene A (chsA) in Petunia hybrida

Chalcone synthase(CHS),coded by the chs 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 88～269 bp,and the sequence 103～201 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),box1 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.