Project description:Mouse BM extracellular vesicles were isolated from Ocn-GFP Topaz mice using the Exoeasy and miRneasy (Qiagen). Small RNA libraries were constructed using NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England biolabs). Libraries were sequenced using Hiseq2500 instrument
Project description:Recently, we demonstrated that RDRs had a general function to synthesize antisense RNAs from sense transcripts of protein-coding genes. In this study, we analyzed whether RDR-mediated antisense RNAs were processed into small RNAs by deep sequencing using SOLiD. Deep sequencing identified 1,645 RDR1/2/6-mediated smRNA loci in drought stress and control conditions.
Project description:Human utilization of the mulberry-silkworm interaction started at least 5,000 years ago and greatly influenced world history through the Silk Road. Complementing the silkworm genome sequence, here we describe the genome of a mulberry species (Morus notabilis C. K. Schneider). In the 330 Mb genome assembly of M. notabilis, we identified 128 Mb of repetitive sequences and 29,338 genes, 60.8% of which were supported by transcriptome sequencing. Mulberry gene sequences appear to evolve ~3 times faster than other Rosales, perhaps facilitating its spread to Europe, Africa, and America. It is among few eudicots but several Rosales not preserving genome duplications in more than 100 million years – however neopolyploid series in mulberry and several others suggest that new duplications may confer benefits. Strikingly, five predicted mulberry miRNAs were found in the hemolymph and silkglands of silkworm, suggesting profound molecular level interactions that promise to expand knowledge of plant-herbivore relationship which constitute key elements of most terrestrial habitats. In addition, we investigated the characters of hemolymph small RNA. small mRNA profiles of silkworm hemolymph in the fifth instar day-5 silkworm were generated by deep sequencing, in twice, using Illumina Hiseq 2000.
Project description:Transposable elements comprise a large proportion of animal genomes. Transcripts of transposable elements are a source for the synthesis of endogenous siRNAs and piRNAs. In order to determine if small RNAs mapped to expressed Tc1-like elements are present during early Xenopus tropicalis development, we used Illumina (Solexa) to sequence small RNAs from gastrula-stage embryos. We obtained about 17 million reads that mapped perfectly to the genome. Small RNAs mapped to selected transposable elements were characterized and the expression of selected small RNAs was experimentally verified during development. This is the first deep sequencing experiment for small RNAs in the Xenopus tropicalis gastrula.
Project description:Transposable elements comprise a large proportion of animal genomes. Transcripts of transposable elements are a source for the synthesis of endogenous siRNAs and piRNAs. In order to determine if small RNAs mapped to expressed Tc1-like elements are present during early Xenopus tropicalis development, we used Illumina (Solexa) to sequence small RNAs from gastrula-stage embryos. We obtained about 17 million reads that mapped perfectly to the genome. Small RNAs mapped to selected transposable elements were characterized and the expression of selected small RNAs was experimentally verified during development. This is the first deep sequencing experiment for small RNAs in the Xenopus tropicalis gastrula. Analysis of small RNAs expressed in the Xenopus tropicalis gastrula.
Project description:We profiled the expression of circulating microRNAs (miRNAs) in mice exposed to gram-positive and gram-negative bacteria using Illumina small RNA deep sequencing. Recombinant-specific gram-negative pathogen Escherichia coli (Xen14) and gram-positive pathogen Staphylococcus aureus (Xen29) were used to induce bacterial infection in mice at a concentration of 1 × 108 bacteria/100 μL of phosphate buffered saline (PBS). Small RNA libraries generated from the serum of mice after exposure to PBS, Xen14, Xen29, and Xen14+Xen29 via the routes of subcutaneous injection (I), cut wound (C), or under grafted skin (S) were analyzed using an Illumina HiSeq2000 Sequencer. Following exposure to gram-negative bacteria alone, no differentially expressed miRNA was found in the injection, cut, or skin graft models. Exposure to mixed bacteria induced a similar expression pattern of the circulating miRNAs to that induced by gram-positive bacterial infection. Upon gram-positive bacterial infection, 9 miRNAs (mir-193b-3p, mir-133a-1-3p, mir-133a-2-3p, mir-133a-1-5p, mir-133b-3p, mir-434-3p, mir-127-3p, mir-676-3p, mir-215-5p) showed upregulation greater than 4-fold with a p-value < 0.01. Among them, mir-193b-3p, mir-133a-1-3p, and mir-133a-2-3p presented the most common miRNA targets expressed in the mice exposed to gram-positive bacterial infection.
Project description:Plant microRNAs (miRNAs) have emerged as important regulators in developmental processes and stress responses in plants. To identify the wound-responsive miRNAs in the leaves of sweet potato, small RNA deep sequencing was conducted on unwounded and wounded leaves (30 min). Total RNAs were isolated for library construction and analyzed by RNA-sequencing via Illumina Genome Analyzer IIx platform. About 16 million total reads were obtained for each sample.
Project description:RNA deep sequencing efforts have revealed abundant expression of small RNAs derived from small nucleolar (sno) RNAs. We employ here spatial RNA deep sequencing to assess the expression of 10-40 nt small RNAs in subcellular compartments of HeLa cells. Total cellular, cytoplasmic, nuclear and nucleolar fractions were isolated, RNA was purified and size-fractionated in a two-step process to yield 10-40 nt RNA fractions. cDNA libraries were constructed and sequenced on a Ion Torrent platform. Vast majority of cellular, cytoplasmic and nuclear small RNA reads were identified as miRNAs. We found the expression of eleven ten miRNAs in the nucleolar preparations using a cut-off rate of 10 reads. Several miRNAs had a greater relative abundance in the nucleolus compared to the other compartments. The nucleolar small RNAs had a unique size distribution consisting of 19-20 and 25 nt RNAs, which were predominantly composed of small snoRNA-derived box C/D RNAs (termed as sdRNA). Sequences from 47 sdRNAs were identified, which mapped both 5M-bM-^@M-^Y and 3M-bM-^@M-^Y ends of the snoRNAs, and retained conserved box C or D motifs. SdRNA reads from SNORD44 comprised 74% of all nucleolar sdRNAs, and were confirmed by Northern blotting as 20 and 25 nt RNAs. This study reveals a rich representation of cell-compartment specific expression of small RNAs and the distinctive unique composition of the nucleolar small RNAs. Examination of small RNAs in cellular subcompartments