Codon usage bias (CUB) phenomenon varies with the species and even within the genes of the same species, where few codons are preferred more frequently than their other synonymous codons. It also categorizes the differences between species. Nucleotide compositional analysis reveals the molecular mechanisms of genes and the evolutionary relationship of a gene in dissimilar plant species. In the present study, three orthologous sequences of each FAE1 (FAE1.1, FAE1.2, and FAE1.3) and FAD2 (FAD2.1, FAD2.2, and FAD2.3) genes, from six Brassica species were accessed using the GenBank database. Further, CUB-related parameters such as nucleotide composition (AT and GC content), relative synonymous codon usage (RSCU), the effective number of codons (ENC), frequency of optimal codons (Fop), relative codon usage bias (RCBS), neutrality plot (GC12 vs. GC3), parity rule-2 [(A3/(A3 + T3) vs. (G3/(G3 + C3)], and correspondence analysis (COA) were analyzed to compare codon bias in U’s triangle Brassica species. The FAE1 genes were AT-biased and FAD2 genes were GC-biased across the studied Brassica species. RSCU values indicated that both the genes had moderate codon usage frequency for selected amino acids. The evolutionary study confirmed that codon usage preference is similar within the species grouped into the same cluster for FAE1; however, B. nigra performed differently for FAD2.2 orthologue. The high ENC value, low Fop, and RSCU value highlighted that FAE1 and FAD2 genes had a low level of gene expression and moderate preference for codon usage across the Brassicas. In addition, neutrality plot, parity rule, and correspondence analysis revealed that natural selection pressure had significantly contributed to CUB for FAE1 genes, whereas mutation and selection pressure occurred for FAD2 genes. This study would help to decode codon optimization, improve the level of expression of exogenous genes, and transgenic engineering to increase fatty acid profiling for the betterment of seed oil in Brassica species.

Erysiphe cruciferarum, a causative agent of powdery mildew disease, has emerged as a serious threat in Brassica juncea and its closely related species. To date, no resistant cultivars have been identified in Brassica species against powdery mildew. Here, we used histopathological, biochemical, and molecular approaches to elucidate the powdery mildew disease progression and host responses in three Brassica cenospecies, namely B. juncea, Camelina sativa, and Sinapis alba. Based on the results of disease progression, S. alba was found to be extremely resistant to powdery mildew infection, whereas B. juncea and C. sativa were highly vulnerable. In addition, the disease spread rate to uninfected parts was comparatively higher in B. juncea and C. sativa. Histopathological results revealed more pathogen-induced cell death in B. juncea and C. sativa compared to S. alba. We also examined the role of antioxidant enzymes such as superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) in B. juncea, C. sativa, and S. alba after powdery mildew infection. Based on our findings, the enzyme activity of SOD, POD, and CAT was relatively higher in S. alba then that of B. juncea and C. sativa after powdery mildew infection. Furthermore, we evaluated the expression levels of salicylic acid (SA) signature genes, including pathogenesis-related protein viz., PR1, PR2, and PR5 in B. juncea, C. sativa, and S. alba after E. cruciferarum infection. Based on our findings, the expression levels of SA marker genes PR1, PR2, and PR5 increased in all three species after infection. However, the fold change was relatively higher in S. alba than in B. juncea and C. sativa. In future, further studies are required to identify the potential candidates in S. alba that are involved in powdery mildew disease resistance.