Carotenoid analysis of sweetpotato Ipomoea batatas and functional identification of its lycopene β- and ε-cyclase genes.

Sweetpotato Ipomoea batatas is known as a hexaploid species. Here, we analyzed carotenoids contained in the leaves and tubers of sweetpotato cultivars 'White Star' (WS) and W71. These cultivars were found to contain several carotenoids unique to sweetpotato tubers such as β-carotene-5,6,5',8'-diepoxide and β-carotene-5,8-epoxide. Next, we isolated two kinds of carotene cyclase genes that encode lycopene β- and ε-cyclases from the WS and W71 leaves, by RT-PCR and subsequent RACE. Two and three lycopene β-cyclase gene sequences were, respectively, isolated from WS, named IbLCYb1, 2, and from W71, IbLCYb3, 4, 5. Meanwhile, only a single lycopene ε-cyclase gene sequence, designated IbLCYe, was isolated from both WS and W71. These genes were separately introduced into a lycopene-synthesizing Escherichia coli transformed with the Pantoea ananatis crtE, crtB and crtI genes, followed by HPLC analysis. β-Carotene was detected in E. coli cells that carried IbLCYb1-4, indicating that the IbLCYb1-4 genes encode lycopene β-cyclase. Meanwhile, the introduction of IbLCYe into the lycopene-synthesizing E. coli led to efficient production of δ-carotene with a monocyclic ε-ring, providing evidence that the IbLCYe gene codes for lycopene ε-(mono)cyclase. Expression of the β- and ε-cyclase genes was analyzed as well.