Baraja-Fonseca, V. | Search bioRxiv

bioRxiv2024

Benchmarking of Low Coverage Sequencing Workflows for Precision Genotyping in Eggplant

Low-coverage whole-genome sequencing (lcWGS) presents a cost-effective solution for genotyping, particularly in applications requiring high marker density and reduced costs. In this study, we evaluated lcWGS for eggplant genotyping using eight founder accessions from the first eggplant MAGIC population (MEGGIC), testing various sequencing coverages and minimum depth of coverage (DP) thresholds with two SNP callers, Freebayes and GATK. Reference SNP panels were used to estimate the percentage of common biallelic SNPs (i.e, true positives, TP) relative to the low coverage datasets (accuracy) and the SNP panels themselves (sensitivity), along with the percentage of TP with the same genotype across the two datasets (genotypic concordance). Sequencing coverages as low as 1X and 2X achieved high accuracy but lacked sufficient sensitivity and genotypic concordance. However, 3X sequencing reached approximately 10% less sensitivity than 5X while maintaining genotypic concordance above 90% at any DP threshold. Freebayes outperformed GATK in terms of sensitivity and genotypic concordance. Therefore, we used this software to conduct a pilot test with some MEGGIC lines from the fifth generation of selfing (S5), comparing their datasets with a gold standard (GS). Sequencing coverages as low as 1X identified a substantial number of TP, with 3X significantly increasing the yield, particularly at moderate DP thresholds. Additionally, at least 30% of the TP were consistently genotyped in all lines when using coverages greater than 2X, regardless of the DP threshold applied. This study highlights the importance of using a GS to reduce false positives and demonstrates that lcWGS, with proper filtering, is a valuable alternative to high-coverage sequencing for eggplant genotyping, with potential applications to other crops.

genomics↗

bioRxiv2023

The irregular fruit green netting: An eggplant domestication trait controlled by the SmGLK2 gene with implications in fruit colour diversification

The distribution of chlorophylls in the eggplant (Solanum melongena) fruit peel can be uniform or display an irregular green netting pattern. The fruit green netting phenotype, manifested as a gradient of dark green netting, more intense in the proximal part of the fruit on a pale green background, is commonly present in eggplant wild relatives as well as in some eggplant landraces. During domestication and modern breeding of eggplant, uniform fruit colour has been selected. However, the fruit green netting contributes to a greater diversity of fruit colours. Here, we have used over 2,300 individuals from several germplasm and experimental populations, including a multi-parental MAGIC population for candidate genomic region identification, an F2 population for BSA-Seq, and advanced backcrosses for edges-to-core fine mapping, to determine that SmGLK2 is the gene underlying the irregular netting in eggplant fruits. We have also analysed the gene sequence of 178 S. melongena accessions and 22 wild relative species for tracing the evolutionary changes that the gene has undergone over the course of domestication. Three different mutations were identified leading to the absence of netting. The main causative indel results in the appearance of a premature stop codon disrupting the protein conformation and function, which was confirmed by Western blotting analysis and confocal microscopy observations. SmGLK2 has a major role in the regulation of chlorophyll biosynthesis in eggplant fruit peel, and therefore in eggplant fruit photosynthesis.

genetics↗