Genome-wide identification and characterization of the RNAi gene families in Brassica rapa L. highlighting their regulatory components and underlying functions involving crop improvement

Brassica rapa is an oilseed crop used for vegetable oil production, with applications in both human consumption and industrial purposes. RNAi-guided gene families play crucial roles in plant growth and development by regulating gene expression in response to various pathogens and abiotic stressors. However, these gene families in this species have not yet been studied in detail. Therefore, this research aims to explore the major RNAi genes, including their characterization and underlying functions for crop improvement. Our analysis identified 4 BrDCL, 13 BrAGO, and 6 BrRDR genes from the Brassica rapa genome. Phylogenetic analysis revealed that the identified genes were evolutionarily related to the RNAi genes in Arabidopsis . The domain, exon-intron, and motif structures of genes and proteins were very similar to those of A. thaliana. Moreover, sequence logo and relative frequency analysis showed that lysine (K), serine(S), valine (V), leucine (L), and glutamic acid (E) were highly significant ( p < 0.01) amino acids in motifs. The genes were distributed across the 10 chromosomes, and segmental duplication was observed in BrAGO1 and BrAGO4. GO analysis showed that genes were involved in important biological processes such as defense response to virus ( p < 0.01), post-transcriptional gene silencing ( p < 0.01), etc., and molecular pathways, for example, RNA polymerase activity ( p < 0.01), siRNA binding( p < 0.01), etc. The largest number of genes was associated with the nucleus, chloroplast, and cytosol. Trans-regulatory analysis showed that the top-ranked TF families accounted for 393 (78%) of 502 regulators, including Dof 115 (23%), bZIP 58(11.55%), C2H2 49(9.76%), ERF 30(6%), BBR-BPC 29(5.77%), MICK-MADS 23(4.58%), MYB, TCP, and WRKY each 23 (4.58%), with AP2 regulating 21(4.18%) TFs. Gene-TF network analysis showed that eight key TF families were highly connected to RNAi genes. The associated cis -acting elements were classified as hormone, light, and various stress-responsive. 10 key RNAi genes interacted with most of the TFs, comprising 302 TFs (~61%), among which BrAGO1a (48 TFs~10%) and BrAGO1b (40 TFs~8%) were influential. Expression analysis demonstrated that most genes showed expression in the root, seeds, flowers, leaves of seedlings, silique, and seed coats. Overall results would provide valuable resources for improving stress-tolerant transgenic lines by initiating knockdown of negative regulators of stress responses, increasing defense-related gene expression, or generating host-induced gene-silencing molecules that target pathogens and pests. Also, by manipulating BrRNAi genes, one can significantly improve crop traits such as disease resistance, abiotic stress tolerance, and crop yield with quality in B. rapa .