Browsing by Author "Dai, Xinbin"

Filter results by typing the first few letters
Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Comprehensive analysis of small RNA-seq data reveals that combination of miRNA with its isomiRs increase the accuracy of target prediction in Arabidopsis thaliana
    (Taylor & Francis Group, 2014) Ahmed, Firoz; Senthil-Kumar, Muthappa; Lee, Seonghee; Dai, Xinbin; Mysore, Kirankumar S; Zhao, Patrick Xuechun
    Along with the canonical miRNA, distinct miRNA-like sequences called sibling miRNAs (sib-miRs) are generated from the same pre-miRNA. Among them, isomeric sequences featuring slight variations at the terminals, relative to the canonical miRNA, constitute a pool of isomeric sibling miRNAs (isomiRs). Despite the high prevalence of isomiRs in eukaryotes, their features and relevance remain elusive. In this study, we performed a comprehensive analysis of mature precursor miRNA (pre-miRNA) sequences from Arabidopsis to understand their features and regulatory targets. The influence of isomiR terminal heterogeneity in target binding was examined comprehensively. Our comprehensive analyses suggested a novel computational strategy that utilizes miRNA and its isomiRs to enhance the accuracy of their regulatory target prediction in Arabidopsis. A few targets are shared by several members of isomiRs; however, this phenomenon was not typical. Gene Ontology (GO) enrichment analysis showed that commonly targeted mRNAs were enriched for certain GO terms. Moreover, comparison of these commonly targeted genes with validated targets from published data demonstrated that the validated targets are bound by most isomiRs and not only the canonical miRNA. Furthermore, the biological role of isomiRs in target cleavage was supported by degradome data. Incorporating this finding, we predicted potential target genes of several miRNAs and confirmed them by experimental assays. This study proposes a novel strategy to improve the accuracy of predicting miRNA targets through combined use of miRNA with its isomiRs.
  • Thumbnail Image
    Item
    pssRNAit-a web server for designing effective and specific plant siRNAs with genome-wide off-target assessment
    (American Society of Plant Biologists, 2020) Ahmed, Firoz; Senthil-Kumar, Muthappa; Dai, Xinbin; Ramu, Vemanna S.; Lee, Seonghee; Mysore, Kirankumar S; Zhao, Patrick Xuechun
    We report an advanced web server named pssRNAit (plant specific small non-coding RNAi tool) that can be used to design a pool of small interfering RNAs (siRNAs) for highly effective, specific, and non-toxic gene silencing in plants. In developing this tool, we integrated the transcript dataset of plants, several rules governing gene silencing, and a series of computational models of the biological mechanism of the RNA interference (RNAi) pathway. The designed pool of siRNAs can be used to construct a long double-stranded RNA (long-dsRNA) and expressed through virus-induced gene silencing (VIGS) or synthetic trans-acting siRNA (syn-tasiRNA) vectors for gene silencing. We demonstrated the performance of pssRNAit by designing and expressing the VIGS constructs to silence Phytoene desaturase (PDS) or a ribosomal protein-encoding gene, RPL10 (QM), in Nicotiana benthamiana. We analyzed the expression levels of predicted intended target and off-target genes using RT-qPCR. We further conducted an RNA-seq-based transcriptome analysis to assess genome-wide off-target gene silencing triggered by the fragments that were designed by pssRNAit, targeting different homologous regions of the PDS gene. Our analyses confirmed the high accuracy of siRNA constructs designed using pssRNAit. The pssRNAit server, freely available at https://plantgrn.noble.org/pssRNAit/, supports the design of highly effective and specific RNAi, VIGS, or syn-tasiRNA constructs for high-throughput functional genomics and trait improvement in more than 160 plant species.