Genetic analysis of carotenoid biosynthesis in chickpea (Cicer arietinum L.) seeds

Abstract

Vitamin A deficiency is a worldwide problem especially in the third world countries. Improvement of carotenoid levels in the edible parts of the crops has been one of the major objectives of many plant breeding programs. Chickpea (Cicer arietinum L.) is an important source of carotenoids. The availability of diverse germplasm resources along with the availability of its genome sequence, makes chickpea an ideal object for studying carotenogenesis in pulses. The objectives of this research were: 1) to identify the genomic regions associated with carotenoid concentration in diverse chickpea accessions and in populations derived from biparental crosses, 2) to examine the effects of environment on carotenogenesis, 3) to examine the relationship between cotyledon colour and carotenoid concentration, and 4) to examine the expression patterns of the genes involved in carotenoid biosynthesis during seed development. A genotypic panel of 172 chickpea accessions was evaluated in 2015 and 2016 with one location per year in Saskatchewan, Canada. The effects of genotype and environment were significant on the concentration of each carotenoid component. The mean and range for the concentration of each component based on the average of two-years data are as follows: 10.14 and 3.5-28.2 µg g-1 for lutein, 0.37 and 0-3.04 µg g-1 for violaxanthin, 1.65 and 0.27-2.84 µg g-1 for zeaxanthin, 0.09 and 0-2.5 µg g-1 for β-carotene respectively. The chickpea genotypic panel consisted of two major subpopulations, kabuli and desi groups, along with an admixture group. Genome-wide association analysis revealed that the genes in the primary steps of carotenoid biosynthesis and those involved in apo-carotenoid production had significant associations with carotenoid concentration in chickpea. Three F2 populations derived from crossing cultivars with green and yellow cotyledons were used to identify QTL associated with carotenoids. Five to eight QTLs responsible for different carotenoid components were identified in each population. In all three populations, the highest phenotypic variation explained by QTL was found for the β-carotene concentration. Cotyledon colour (CotCol) was mapped on linkage group 8 in each population. A positive and significant relationship between cotyledon colour and carotenoid concentration was identified in this experiment. The structure, genomic location, and copy number of 29 genes involved in carotenoid and isoprenoid pathways were retrieved in the chickpea genome. Two missense mutations were found in zeta carotene isomerase (ZISO2) in CDC Verano, a green cotyledon kabuli cultivar, which might explain the higher carotenoid concentration in this cultivar. The expression patterns of 19 genes from the carotenoid pathway were analyzed in five chickpea cultivars at different seed developmental stages. The highest expression level from all the genes was observed at eight and 16 days post-anthesis across the five cultivars. The highest carotenoid concentration and expression levels of the carotenoid genes were found in CDC Jade, a desi cultivar with green cotyledons. Based on gene expression analysis, the desaturation and isomerisation reactions positively affected the carotenoid concentration, while hydroxylation adversely affected the carotenoid concentration. The results from this research could help breeders to develop chickpea cultivars with improved carotenoid/provitamin A levels through molecular breeding.