Department of Plant Production and Forestry Science, University of Lleida-Agrotecnio CERCA Center, Lleida, Spain; School of Life Sciences, Changchun Normal University, Changchun, China.
Beijing Key Laboratory of Captive Wildlife Technologies, Beijing Zoo, Beijing, China.
Methods Enzymol. 2022;671:511-526. doi: 10.1016/bs.mie.2021.09.016. Epub 2021 Oct 25.
Carotenoids are a large class of important lipid-soluble phytonutrients that are widely used as nutritional supplements due to their health-promoting activities. For example, β-carotene is the precursor for vitamin A synthesis, and astaxanthin is a powerful antioxidant. However, these carotenoids cannot be synthesized de novo by humans. These properties of β-carotene and astaxanthin make them attractive targets for metabolic engineering in rice (Oryza sativa) endosperm because rice is an important staple food in developing countries, and rice endosperm is devoid of carotenoids. In this chapter, we introduce an assay based on rice embryogenic callus for the rapid functional characterization of genes involved in carotenoid biosynthesis and accumulation. The system is also an ideal platform to characterize cereal endosperm specific promoters. Four diverse cereal endosperm specific promoters were demonstrated to be active in rice callus despite their restricted activity in mature plants. The use of endosperm specific promoters that are expressed in rice callus, but remain silent in regenerated vegetative tissue, directs accumulation of carotenoids in the endosperm without interfering with plant growth. Rice callus is a useful platform for improving gene editing methods and for further optimizing pathway engineering. Thus, the rice callus platform provides a unique opportunity to test strategies for metabolic engineering of synthetic carotenoid pathways, leading to novel carotenoid-biofortified crops.
类胡萝卜素是一大类重要的脂溶性植物营养素，由于其具有促进健康的活性，因此被广泛用作营养补充剂。例如，β-胡萝卜素是维生素 A 合成的前体，虾青素是一种强大的抗氧化剂。然而，这些类胡萝卜素不能被人体从头合成。β-胡萝卜素和虾青素的这些特性使得它们成为水稻（Oryza sativa）胚乳代谢工程的有吸引力的目标，因为水稻是发展中国家的重要主食，而水稻胚乳中不含类胡萝卜素。在本章中，我们介绍了一种基于水稻胚性愈伤组织的快速功能鉴定方法，用于鉴定参与类胡萝卜素生物合成和积累的基因。该系统也是鉴定谷物胚乳特异性启动子的理想平台。尽管在成熟植物中活性受限，但四个不同的谷物胚乳特异性启动子在水稻愈伤组织中表现出活性。使用在水稻愈伤组织中表达但在再生营养组织中沉默的胚乳特异性启动子，可将类胡萝卜素积累定向到胚乳中，而不干扰植物生长。水稻愈伤组织是改进基因编辑方法和进一步优化途径工程的有用平台。因此，水稻愈伤组织平台为测试合成类胡萝卜素途径代谢工程的策略提供了独特的机会，从而产生了新型富含类胡萝卜素的作物。
