Validation of KASP-SNP markers in cassava germplasm for marker-assisted selection of increased carotenoid content and dry matter content

Abstract

Provitamin A biofortification and increased dry matter content are important breeding targets in cassava improvement programs worldwide. Biofortified varieties contribute to the alleviation of provitamin A deficiency, a leading cause of preventable blindness common among pre-school children and pregnant women in developing countries particularly Africa. Dry matter content is a major component of dry yield and thus underlies overall variety performance and acceptability by growers, processors, and consumers. Single nucleotide polymorphism (SNP) markers linked to these traits have recently been discovered through several genome-wide association studies but have not been deployed for routine marker-assisted selection (MAS). This is due to the lack of useful information on markers’ performances in diverse genetic backgrounds. To overcome this bottleneck, technical and biological validation of the loci associated with increased carotenoid content and dry matter content were carried out using populations independent of the marker discovery population. In the present study, seven previously identified markers for these traits were converted to a robust set of uniplex allele-specific polymerase chain reaction (PCR) assays and validated in two independent pre-breeding and breeding populations. These assays were efficient in discriminating marker genotypic classes and had an average call rate greater than 98%. A high correlation was observed between the predicted and observed carotenoid content as inferred by root yellowness intensity in the breeding (r = 0.92) and pre-breeding (r = 0.95) populations. On the other hand, dry matter content-markers had moderately low predictive accuracy in both populations (r< 0.40) due to the more quantitative nature of the trait. This work confirmed the markers’ effectiveness in multiple backgrounds, therefore, further strengthening their value in cassava biofortification to ensure nutritional security as well as dry matter content productivity. Our study provides a framework to guide future marker validation, thus leading to the more routine use of markers in MAS in cassava improvement programs.

SNP-based marker-assisted selection for high provitamin A content in African cassava genetic background

“…However, the best model chosen using a stepwise regression method returned a simple model only fitted with the most significant predictor at the phytoene synthase gene indicating the elimination of other non-significant markers in the prediction model for TCC and root colour. The low phenotypic variation of TCC and root colour explained by the three markers S1_30543962, S5_3387558 and S8_25598183 suggests that the newly uncovered QTLs for pVAC have a minor effect on the trait, as previously reported by Ige et al 35 The best prediction model fitted with the most significant marker was validated using a bootstrap resampling strategy. The correlation coefficient between the predicted and the observed TCC values ranged from 0.36 to 0.80 with an average of 0.63 (Figure 6) whereas a correlation ranging from 0.66 to 0.89 with a mean of 0.75 was found between predicted and observed root colour score (Figure 7).…”

“…A similar study using different cassava populations resulted in a higher accuracy of 80%. 35 The relatively lower prediction accuracy from our study could be explained by the small population size derived from five parents and the narrow range of variation in root pigmentation (between cream and yellow) after discarding white root clones at the early stages of selection.…”

“…The KASP genotyping was performed as described by Codjia et al 41 Four top SNPs (S1_24155522, S1_30543962, S5_3387558 and S8_25598183) coming from phytoene synthase 2 on chromosome 1 and new loci on chromosomes 1, 5 and 8 extracted from Rabbi et al 33 were used for screening the cassava genotypes for pVAC. The pVAClinked markers successfully converted to allele-specific PCR assay 35 enabled the molecular screening. Yellow variety IITA-IBA-TMS 07/0593 (one of the female parents) and white variety IITA-IBA-TMS 30555 were used as positive and negative controls, respectively, in the screening for carotene content.…”

“…33 Four of the genome-wide uncovered markers (S1_24155522, S1_30543962, S5_3387558, and S8_25598183) linked to pVAC were recently converted to allele-specific PCR assays to facilitate their routine use in MAS. 35 We carried out MAS using these SNP markers on a pVAC breeding population developed at the University of Ibadan, Nigeria. We present the performance of the markers in predicting total carotenoid content and root colour score.…”

Genetic parameters considering traits of importance for cassava biofortification

“…Dry matter content (DMC; %) was determined with the oven drying method (Ige et al 2022). The moisture from the roots (sample of 100 g) was obtained after drying in forced air circulation oven (Dehydrator Pardal PE60, Petrópolis, Brazil) at 60 °C to constant weight.…”