Construction And Genetic Transformation Of TaHYD Overexpression And RNA Interference Vectors In Wheat

Wheat is one of the most important food crops worldwide. Improving the nutritional value of wheat is an important subject in wheat genetic breeding. Carotenoid content is one of the important parameters to judge the nutritional value. Carotenoid is a main resource of provitamin A. However, the carotenoid and provitamin A content is very low in the grains of common wheat.We know little about carotenoid synthesis and only few reports had reported about improving carotenoid through genetic engineering. Cloning and identification of more genes in carotenoid metabolic pathway is important for improving the nutritional value of wheat.Carotene hydroxylase(HYD) is a key enzyme in carotenoid metabolic pathway of plants by converting β-carotene into Xanthophyll. In the present study, a novel carotene hydroxylase gene, designed TaHYD, was cloned from wheat by homology cloning strategy. Then TaHYD overexpression and RNA interference vectors were constructed and they were transformed into wheat mode variety Bobwhite via particle bombardment to study the function of HYD in carotenoid biosynthetic pathway. After extracting the DNA of transgenic wheat, 2 positive overexpression lines and 6 positive RNA interference lines were confirmed by Polymerase Chain Reaction(PCR). The results of qRT-PCR and High Performance Liquid Chromatography indicate that the expression of TaHYD in overexpression lines were up-regulated accompanied with no significant changes of carotenoid content, while the expression of TaHYD in RNA interference lines were down-regulated accompanying with significant increase in the β-carotene content. Our results not only demonstrate that TaHYD is bound up with the content of β-carotene in wheat endosperm, but also provide theoretical basis and experimental evidence for improving β-carotene content in wheat endosperm by genetic engineering methods.