Genome-wide profiling of miRNAs and other small non-coding RNAs in the Verticillium dahliae inoculated cotton roots
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ABSTRACT: MicroRNAs (miRNAs) and small interfering RNAs (siRNAs) are short (19â25 nucleotides) non-coding RNA molecules that have large-scale regulatory effects on development and on stress responses in plants.The objective of this study is to investigate the transcriptional profile of miRNAs and other small non-coding RNAs in Verticilliumâinoculated cotton roots. Four small RNA libraries were constructed from mocked and infected roots of two cotton cultured species which are with different Verticillium tolerance (âHai-7124â, Gossypium barbadense L., a Verticillium-tolerant cultivar, and âYi-11â, Gossypium hirsutum L. a Verticillium-sensitive cultivar). The length distribution of obtained small RNA pools was significantly different among libraries. A total of 215 conserved miRNA families were identified in the two cotton species, of them 14 are novel. There were >65 families with different expression between two libraries. We also identified two ta-siRNAs and thousands of endogenous siRNA candidates, and hundred of them exhibited altered expression after inoculation of Verticillium. The profiling of these miRNAs and other small non-coding RNAs lay the foundation for further understanding of small RNAs function in the regulation of Verticillium defence responses in cotton roots. Examination of 2 different traetments in 2 cotton types.
Project description:MicroRNAs (miRNAs) and small interfering RNAs (siRNAs) are short (19â25 nucleotides) non-coding RNA molecules that have large-scale regulatory effects on development and on stress responses in plants.The objective of this study is to investigate the transcriptional profile of miRNAs and other small non-coding RNAs in Verticilliumâinoculated cotton roots. Four small RNA libraries were constructed from mocked and infected roots of two cotton cultured species which are with different Verticillium tolerance (âHai-7124â, Gossypium barbadense L., a Verticillium-tolerant cultivar, and âYi-11â, Gossypium hirsutum L. a Verticillium-sensitive cultivar). The length distribution of obtained small RNA pools was significantly different among libraries. A total of 215 conserved miRNA families were identified in the two cotton species, of them 14 are novel. There were >65 families with different expression between two libraries. We also identified two ta-siRNAs and thousands of endogenous siRNA candidates, and hundred of them exhibited altered expression after inoculation of Verticillium. The profiling of these miRNAs and other small non-coding RNAs lay the foundation for further understanding of small RNAs function in the regulation of Verticillium defence responses in cotton roots.
Project description:Genetic male sterility (GMS) in cotton (Gossypium hirsutum) plays an important role in the utilization of hybrid vigor. However, the molecular mechanism of the GMS is still unclear. While numerous studies have demonstrated that microRNAs (miRNA) regulate flower and anther development, whether different small RNA regulations exist in GMS and its wild type is unclear. To investigate the global expression and complexity of small RNAs during cotton anther development, three small RNA libraries were constructed from the anthers of three development stages each from fertile wild type (WT) and its GMS mutant cotton.
Project description:Genetic male sterility (GMS) in cotton (Gossypium hirsutum) plays an important role in the utilization of hybrid vigor. However, the molecular mechanism of the GMS is still unclear. While numerous studies have demonstrated that microRNAs (miRNA) regulate flower and anther development, whether different small RNA regulations exist in GMS and its wild type is unclear. To investigate the global expression and complexity of small RNAs during cotton anther development, three small RNA libraries were constructed from the anthers of three development stages each from fertile wild type (WT) and its GMS mutant cotton. Examination of different miRNA profiles in 2 lines.
Project description:Cotton is the main source of natural fiber in the textile industry, making it one of the most economically important fiber crops in the world. Verticillium wilt, caused by the pathogenic fungus Verticillium dahlia, is one of the most damaging biotic factors limiting cotton production. Mechanistic details of cotton defense responses to verticillium wilt remain unclear. In this study, GFP-labeled strain of V. dahlia was used to track colonization in cotton roots, and clear conidial germination could be observed at 48 hours post-inoculation (hpi), marking this as a crucial time point during infection. Transcriptome analysis identified 1,523 and 8,270 differentially expressed genes (DEGs) at 24 hpi and 48 hpi, respectively. Metabolomic screening found 78 differentially accumulated metabolites (DAMs) at 48 hpi. Conjoint analysis indicated that the phenylpropanoid biosynthesis pathway was activated in cotton infected with V. dahliae. The five metabolites in the phenylpropanoid biosynthesis pathway, including caffeic acid, coniferyl alcohol, coniferin, scopoletin and scopolin, could significantly inhibit V. dahlia growth in vitro, implicating their roles in cotton resistance to Verticillium wilt. The findings expand our understanding of molecular mechanisms underlying the pathogen defense response against V. dahlia infection in upland cotton, which may lead to future insights into controlling Verticillium wilt disease.
Project description:To dissect the roles of miRNAs in fiber development, we sequenced small RNAs from ovules -1 to +1 day post anthesis (DPA) and young leaves. A series of conserved and novel miRNA were identified from cotton EST database and the genome of G.raimondii, many of which were shown to be expressed differentially between ovule and leaf, indicating their potential roles in fiber development or leaf development. Cotton (Gossypium Hirsutum L.) cultivar TM-1 were grown in the greenhouse. Cotton ovules -1 to +1 day post anthesis (DPA) and young leaves were harvested for five biological replicates and immediately frozen in liquid nitrogen. They were then stored at -80M-BM-0C following RNA extraction. Totals RNAs were extracted from each tissue sample using the mirVana miRNA isolation kit (Ambion, Austin, TX) according to the manufacturerM-bM-^@M-^Ys protocol. The small RNA samples extracted from the five biological replicates were pooled together for leaf and ovule, respectively. Finally, the construction of pooled small RNA libraries and sequenced were performed by LC Sciences (Houston, TX) using Illumina high-throughput sequencing platform.
Project description:Genomic approaches to the discovery of promoters for sustained expression in cotton (Gossypium hirsutum L.) under field conditions: expression analysis in transgenic cotton and Arabidopsis of a Rubisco small subunit promoter identified using EST sequence analysis and cDNA microarrays. Keywords: Promoter Discovery
Project description:Verticillium wilt which is caused by Verticillium dahliae causes massive annual losses of cotton yield. Control by conventional mechanisms is not possible due to wide host range and longevity of dormant fungi in the soil in case of absence of a suitable host. Plants have developed various mechanisms to boost their immunity against various diseases, and one of which is through the induction of various genes. In this research work, carried out of RNA sequencing and identified the members of the ABC genes are critical in enhancing resistance to V. dahliae infection. A total of 166 ABC genes were identified in Gossypium raimondii with varying physiochemical properties. A novel ABC gene, Gorai.007G244600 was found to be highly upregulated, its homolog in the tetraploid cotton Gh_D11G3432, was then silenced through virus induced gene silencing (VIGS) in tetraploid cotton, the mutant cotton seedlings that have the ability to tolerate V. dahliae infection were significantly reduced. Evaluation of oxidant, hydrogen peroxide (H2O2) and malondialdehyde (MDA) were found to have increased levels in the leaves of the mutant compared to the wild types. In addition, antioxidant enzymes, peroxidase (POD), catalase (CAT) and superoxide dismutase (SOD) concentration levels were significantly reduced in the mutant cotton compared to the wild types. Moreover, expression levels of the biotic stress genes, cotton polyamine oxidase (GhPAO), cotton ribosomal protein L18(GhRPL18) and cotton polygalacturonase-inhibiting protein-1 (GhPGIP1) were all downregulated in the mutant but highly upregulated in the wild cotton tissues. The outcome of this research has shown that ABC genes could be playing an important role in enhancing immunity of cotton to V. dahliae infection and can be explored in developing more resilient cotton genotypes with improved resistance to V. dahliae infection.
Project description:In this study, two kinds of island cotton materials with different resistances of Verticillium dahliae were used for Verticillium dahliae infection, the proteomic was used to detect all the protein expression levels in the roots of the two kinds of cotton, and the spectronaut pulsar x software was selected to analyze the data
Project description:In this study, two small RNA libraries and two degradome libraries were constructed from roots of Al-treated and Al-free Glycine soja seedlings. For miRNA, a total of 7,287,655 and 7,035,914 clean reads in Al-treated and Al-free small RNAs libraries were generated, and 105 known miRNAs ,51 p3/p5 strands of known miRNA and 80 novel miRNAs were identified. Among them, expression of 34 miRNAs was responsive to Al stress. Through degradome sequencing, 82 and 11 genes were identified as tagerts of known and novel miRNAs obtained from this study, respectively. Gene Ontology (GO) annotations of target transcripts indicated that 52 out of 66 targets cleaved by conserved miRNA families may play role in regulation of transcription. sample 1: Examination of small RNA in Al-free wild soybean roots; sampple 2: Examination of small RNA in Al-treated wild soybena roots; sample 3: identification of miRNA targets in Al-free wild soybean roots; sample 4: identification of miRNA targerts in Al-treated wild soybean roots
Project description:Genomic approaches to the discovery of promoters for sustained expression in cotton (Gossypium hirsutum L.) under field conditions: expression analysis in transgenic cotton and Arabidopsis of a Rubisco small subunit promoter identified using EST sequence analysis and cDNA microarrays. RNAs from 8-week old plants were compared to RNAs from 10-, 14-, 20- and 22-week old plants on separate arrays each with a dye swap. The 20 and 22 weeks hybridisations each had a biological replicate (each with a dye swap) as well as a second independent hybridisation (with dye swap) with one of the 22 week RNAs. One of the 22-week slides was of low quality and had to be discarded.