Enhancing beta-carotene content in potato by rnai-mediated silencing of the beta-carotene hydroxylase gene

Abstract

Plant carotenoids are lipid soluble pigments that play key roles in numerous plant functions. They also play significant roles in the human diet by serving as precursors for vitamin A synthesis and by reducing the occurrence of certain diseases. The purpose of this work was to identify novel methods for enhancing betacarotene content in potato, a major staple food crop. In particular, we used RNA interference (RNAi) to silence the beta-carotene hydroxylase gene (bch), which converts beta-carotene to zeaxanthin. Agrobacterium tumefaciens-mediated transformation was employed to introduce two RNAi constructs into three different potato lines (‘Yema de Huevo’, 91E22, and ‘Desiree’). One construct contained the tuber-specific granulebound starch synthase (GBSS) promoter, and the other contained the strong constitutive cauliflower mosaic virus 35S (CaMV 35S) promoter. Eighty-six percent of the silenced lines had altered carotenoid profiles, as revealed by HPLC. Beta-carotene content was increased from trace amounts in wild type tubers up to 331 μg 100 g1 fresh weight. In addition, some transformants exhibited a significant decrease in zeaxanthin content and/or an increase in lutein. In general, transformants derived from the GBSS construct contained more beta-carotene than CaMV 35S transformants. Reverse-transcriptase PCR (RT-PCR) analysis of bch RNA abundance in tubers demonstrated that the extent of bch silencing varied between transformants, and was in most cases associated with the level of beta-carotene. Similarly, RT-PCR showed that bch silencing also occurred in leaves, but primarily in the CaMV 35S lines. These results demonstrate that silencing bch has the potential to increase the content of two health-promoting carotenoids, betacarotene and lutein, in potato.