Molecular Cloning and Expression Analysis of the ζ-Carotene Desaturase Gene in Chinese kale (Brassica oleracea var. alboglabra Bailey)

ζ-Carotene desaturase (ZDS) is an important enzyme in carotenoid biosynthesis. Here, the Brassica oleracea var. alboglabra ZDS (BoaZDS) gene was cloned from Chinese kale via reverse transcription-polymerase chain reaction (RT-PCR) and deposited in GenBank (accession number KY662297). The BoaZDS gene contains an open reading frame of 1 686 bp that encodes a 561-amino acid protein. Sequence analysis indicates that the ZDS protein is apparently conserved during plant evolution and is most closely related to B. oleracea var. capitata and B. rapa. The promoter sequence of the BoaZDS gene was predicted to harbor several cis-acting elements that are related to light and phytohormone responses. Semi-quantitative RT-PCR analysis showed that BoaZDS expression varied among different developmental stages and organs. Relative ZDS expression remained stable during germination and seedling stages and rapidly increased at the mature leaf stage. The leaves showed the highest ZDS expression levels compared to the other organs. ZDS expression decreased in all flower tissues during blooming. The fused protein of BoaZDS was obtained by prokaryotic expression. Heterologous expression of BoaZDS in Escherichia coli confirmed that BoaZDS encodes a functional ζ-carotene desaturase that increases β-carotene accumulation in E. coli cells harboring a β-carotene-producing plasmid. The findings of the present study provide a molecular basis for the elucidation of ZDS gene function in Chinese kale. Keywords: Chinese kale, Brassica oleracea var. alboglabra Bailey, carotenoids, ζ-carotene desaturase (ZDS), cloning, gene expression, prokaryotic expression, heterologous expressio