Cloning and expression analysis of MYB12 in Lilium oriental hybrid ‘Sorbonne’

To reveal the regulatory mechanism of the anthocyanin synthesis pathway in Lilium, we isolated cDNA, gDNA, and the MYB12 gene promoter from the petals of the L. oriental hybrid ‘Sorbonne’. Results showed that the cDNA sequence contained a 720 bp open reading frame (ORF) encoding a 239 amino acid polypeptide with two typical DNA-binding domains. The 863 bp nucleotide sequence was amplified from the extracted DNA and the gene consisted of three exons and two introns. Furthermore, the amino acid sequence had highest similarity with MYB transcription factors in Tulipa fosteriana W. Irving. Phylogenetic analysis showed that the phylogenetic evolution of the transcription factor was in accordance with taxonomic classification, and it could be grouped with genes that regulate anthocyanin biosynthesis in the MYB gene family. The 2143 bp promoter sequence of the LhsorMYB12 gene was obtained using chromosome walking technology. The promoter element prediction results showed the existence of a core promoter element (TATA box), MYB protein binding sites, photoreactive elements, and other regulation motifs (e.g., circadian rhythm response motif). RT-PCR analysis indicated that the expression of LhsorMYB12 could be detected in the tepals, filaments, stigmas, and styles. qRT-PCR analysis showed that the relatively high expression occurred in blooming period (12 cm) tepals, and then began to decrease. However, there was a difference between the initial stage of expression in the inner and outer perianths. Moreover, dark treatment resulted in a reduction in the expression of LhsorMYB12, with the lowest expression found after an hour of treatment. Under light conditions, the expression level of the gene increased at first, then decreased, and then continued to rise with increasing treatment time. The LhsorMYB12 expression pattern results might relate to the corresponding cis-acting elements in its promoter.