Development of a high-throughput phenotyping method for measuring sorghum carotenoids
Date
2024
Journal Title
Journal ISSN
Volume Title
Abstract
Vitamin A deficiency is the leading cause of preventable blindness in young children, and also leads to infertility and decreased immune function. Humans cannot synthesize vitamin A, so it must be consumed in the diet, either as preformed vitamin A or as provitamin carotenoid (PVACs) in plant foods. Access to PVACs is limited in sub-Saharan Africa where many diets are less diverse and primarily consist of cereals, such as sorghum, with intrinsically low concentrations of PVACs. Therefore, biofortification breeding efforts aim to increase PVACs to biologically relevant levels to reduce global vitamin A deficiency. In order to select and breed high carotenoid varieties, thousands of progeny in a breeding program must be phenotyped. High-performance liquid chromatography (HPLC) is the gold standard carotenoid phenotyping method; however, it is expensive and time-consuming, making it impractical for large-scale screening. We hypothesized that a high-throughput phenotyping (HTP) method using UV-VIS spectrophotometry can identify high carotenoid sorghum lines for selection during breeding. In this study, a simple and rapid method for carotenoid extraction and UV-VIS spectrophotometric detection in a 96-well plate format was developed and validated. To develop the HTP method, we measured 60 samples using both HPLC and UV-VIS, identifying a strong correlation (R2=0.62, p-value<4.51x10-14) between total carotenoid concentrations measured with the HTP method and the gold standard HPLC method. To validate the HTP method, we measured carotenoids in 249 lines in a biparental breeding family, using both HPLC and UV-VIS, and again identified a strong correlation (R2=0.61, p-value<2.2x10-16). We also compared the predictability of the UV-VIS method to that of a simple visual inspection of grain color and found that the UV-VIS method performed significantly better. This promising HTP method will enable rapid screening of a large number of samples, helping breeders more efficiently make selections for carotenoid biofortification.