Genetic factors and biochemical pathways regulating post-harvest aflatoxin contamination in cultivated peanut
Korani, Walid Ahmed Abdelmoneim
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Aflatoxin accumulates in peanut seeds in the field (pre-harvest) or during storage (post-harvest) as a result of Aspergillus spp. infection. It is one of the most challenging diseases for peanut as toxin biosynthesis is dependent on environment, which makes identification of resistant genotypes difficult. Therefore, this study was carried out to discover genetic factors and biochemical pathways that underlie resistance to postharvest aflatoxin contamination of cultivated peanut. Since accumulated evidence supports the importance of the lipoxygenase (LOX) gene superfamily in plant defense against many diseases, LOX genes of cultivated peanut were identified. In addition, functional classification, evolutionary analysis, and in-depth expression analysis were carried out. Moreover, the expression responses to different diseases including aflatoxin contamination were estimated. To identify resistant genotypes, a protocol of tracking and assaying the A. flavus infection and the subsequent aflatoxin accumulation was developed using a GFP-expressing strain. In addition, a phenotyping tool was designed, designated SICIA (Seed Infection Coverage and Intensity Analyzer) using a Matlab script. The application of this protocol combined with different statistical models enabled the identification of a highly reliable resource for resistance in cultivated peanut. Moreover, this approach allowed differentiation between the genotype response to A. flavus infection and the ability to accumulate aflatoxins. Genetic mapping of putative resistance genes requires DNA sequence variation. Since cultivated peanut is tetraploid, extracting true SNPs directly from next generation sequencing data using currently available filtering tools is challenging. Therefore, a machine learning tool for refining SNP calling from sequence data of polyploids was designed, designated SNP-ML. RNA-seq analysis was carried out for A. flavus-infected resistant and susceptible peanut genotypes to determine the factors associated with the resistance response for aflatoxin accumulation in cultivated peanut. An R package was designed to conduct KEGG enrichment analysis for polyploids, designated keggseq. The application of this package revealed the importance of alpha-linolenic acid and protein processing in the endoplasmic reticulum in the resistance response. The analysis also included application of different tools for differential expression analysis of time course experiments, expression clustering, GO enrichment analysis, de novo assembly, annotation and co-expression network analysis.