small rna discovery short read sequencing

Despite linkage to chromosome 16q in 1996, the mutation causing spinocerebellar ataxia type 4 (SCA4), a late-onset sensory and cerebellar ataxia, remained unknown. Here, using long-read single-strand whole-genome sequencing (LR-GS), we identified a heterozygous GGC-repeat expansion in a large Utah pedigree encoding polyglycine (polyG) in zinc finger homeobox protein 3 (ZFHX3), also known as AT-binding transcription factor 1 (ATBF1). We queried 6,495 genome sequencing datasets and identified the repeat expansion in seven additional pedigrees...

BACKGROUND: Theaceae, comprising 300 + species, holds significance in biodiversity, economics, and culture, notably including the globally consumed tea plant. Stewartia gemmata, a species of the earliest diverging tribe Stewartieae, is critical to offer insights into Theaceae's origin and evolutionary history. RESULT: We sequenced the complete organelle genomes of Stewartia gemmata using short/long reads sequencing technologies. The chloroplast genome (158,406 bp) exhibited a quadripartite structure including the large single-copy region (LSC), a small single-copy region (SSC), and a pair of inverted repeat regions (IRs); 114 genes encoded 80 proteins, 30 tRNAs, and four rRNAs...

Capturing and sequencing small RNAs is standard practice, however identification of a group of these small RNAs-small interfering RNAs (siRNAs)-has been more difficult. We present smalldisco, a command-line tool for small interfering RNA discovery and annotation from small RNA-seq datasets. smalldisco can distinguish short reads that map antisense to an annotated genomic feature (e.g., exons or mRNAs), annotate these siRNAs, and quantify their abundance. smalldisco also uses the program Tailor to quantify 3' non-templated nucleotides of siRNAs or any small RNA species...

The analysis of the combined mRNA and miRNA content of a biological sample can be of interest for answering several research questions, like biomarkers discovery, or mRNA-miRNA interactions. However, the process is costly and time-consuming, separate libraries need to be prepared and sequenced on different flowcells. Combo-Seq is a library prep kit that allows us to prepare combined mRNA-miRNA libraries starting from very low total RNA. To date, no dedicated bioinformatics method exists for the processing of Combo-Seq data...

Alternative open reading frames (AltORFs) are unannotated sequences in genome that encode novel peptides or proteins named alternative proteins (AltProts). Although ribosome profiling and bioinformatics predict a large number of AltProts, mass spectrometry (MS) as the only direct way of identification, is hampered by the short lengths and relative low abundance of AltProts. There is an urgent need for improvement of MS methodologies for AltProt identification. Here we report an approach based on size-exclusion chromatography (SEC) for simultaneous enrichment and fractionation of AltProts from complex proteome...

Short open reading frames (sORFs) refer to the small nucleic fragments no longer than 303 nt in length that probably encode small peptides. To date, translatable sORFs have been found in both untranslated regions of messenger ribonucleic acids (RNAs; mRNAs) and long non-coding RNAs (lncRNAs), playing vital roles in a myriad of biological processes. As not all sORFs are translated or essentially translatable, it is important to develop a highly accurate computational tool for characterizing the coding potential of sORFs, thereby facilitating discovery of novel functional peptides...

RNA sequencing has become the method of choice to study the transcriptional landscape of phage-infected bacteria. However, short-read RNA sequencing approaches generally fail to capture the primary 5' and 3' boundaries of transcripts, confounding the discovery of key transcription initiation and termination events as well as operon architectures. Yet, the elucidation of these elements is crucial for the understanding of the strategy of transcription regulation during the infection process, which is currently lacking beyond a handful of model phages...

The transcriptomes of Pseudomonas aeruginosa clone C isolates NN2 and SG17M during the mid-exponential and early stationary phases of planktonic growth were evaluated by direct RNA sequencing on the nanopore platform and compared with established short-read cDNA sequencing on the Illumina platform. Fifty to ninety percent of the sense RNAs turned out to be rRNA molecules, followed by similar proportions of mRNA transcripts and noncoding RNAs. The two platforms detected similar proportions of uncharged tRNAs and 29 yet-undescribed antisense tRNAs...

Existing research has shown that there are a large amount of non-coding RNAs (ncRNAs) in organisms. Short open reading frames (sORFs) abundantly exist in molecular sequences inaccurately annotated as ncRNAs. Several sORFs can be transcribed and translated into evolutionarily conserved micropeptides, which were ignored in previous studies due to short sequence lengths and the limitations of research techniques. To date, sORF-encoded micropeptides with various functions have been found to play important roles in regulating vital biological activities...

BACKGROUND: Several studies have mined short-read RNA sequencing datasets to identify long non-coding RNAs (lncRNAs), and others have focused on the function of individual lncRNAs in abiotic stress response. However, our understanding of the complement, function and origin of lncRNAs - and especially transposon derived lncRNAs (TE-lncRNAs) - in response to abiotic stress is still in its infancy. RESULTS: We utilized a dataset of 127 RNA sequencing samples that included total RNA datasets and PacBio fl-cDNA data to discover lncRNAs in maize...

Fish size is a complex trait determined by the interaction of environmental and genetic factors. While evidence exists that fish final length and size are partially determined by muscle fibre structure. However, the molecular basis behind final body size and muscle fibre composition remains yet to be fully understood. Here we use the fish model zebrafish (Danio rerio) to explore the molecular mechanisms involved in muscle fibre number, muscle growth and their role on animal size determination. In order to do so, we used two lines of zebrafish artificially selected by Amaral and Johnston 2012 for body length to obtain large (L) and small (S) genetic lineages...

Next generation sequencing (NGS) technologies are becoming routinely employed in different fields of virus research. Different sequencing platforms and sample preparation approaches, in the laboratories worldwide, contributed to a revolution in detection and discovery of plant viruses and viroids. In this work, we are presenting the comparison of two RNA sequence inputs (small RNAs vs. ribosomal RNA depleted total RNA) for the detection of plant viruses by Illumina sequencing. This comparison includes several viruses, which differ in genome organization and viroids from both known families...

Riboswitches and attenuators are cis-regulatory RNA elements, most of which control bacterial gene expression via metabolite-mediated, premature transcription termination. We developed an unbiased experimental approach for genome-wide discovery of such ribo-regulators in bacteria. We also devised an experimental platform that quantitatively measures the in vivo activity of all such regulators in parallel and enables rapid screening for ribo-regulators that respond to metabolites of choice. Using this approach, we detected numerous antibiotic-responsive ribo-regulators that control antibiotic resistance genes in pathogens and in the human microbiome...

BACKGROUND: Development of sequencing technologies and supporting computation enable discovery of small RNA molecules that previously escaped detection or were ignored due to low count numbers. While the focus in the analysis of small RNA libraries has been primarily on microRNAs (miRNAs), recent studies have reported findings of fragments of transfer RNAs (tRFs) across a range of organisms. RESULTS: Here we describe Drosophila melanogaster tRFs, which appear to have a number of structural and functional features similar to those of miRNAs but are less abundant...

High throughput, cost effective next generation sequencing (NGS) has enabled the publication of genome sequences for Cricetulus griseus and several Chinese hamster ovary (CHO) cell lines. RNA-Seq, the utilization of NGS technology to study the transcriptome, is expanding our understanding of the CHO cell biological system in areas ranging from the analysis of transcription start sites to the discovery of small noncoding RNAs. The analysis of RNA-Seq data, often comprised of several million short reads, presents a considerable challenge...

miRNA is known to regulate up to several hundreds coding genes, thus the integrated analysis of miRNA and mRNA expression data is an important problem. Unfortunately, the integrated analysis is challenging since it needs to consider expression data of two different types, miRNA and mRNA, and target relationship between miRNA and mRNA is not clear, especially when microarray data is used. Fortunately, due to the low sequencing cost, small RNA and RNA sequencing are routinely processed and we may be able to infer regulation relationships between miRNAs and mRNAs more accurately by using sequencing data...

The discovery of novel microRNA (miRNA) and piwi-interacting RNA (piRNA) is an important task for the understanding of many biological processes. Most of the available miRNA and piRNA identification methods are dependent on the availability of the organism's genome sequence and the quality of its annotation. Therefore, an efficient prediction method based solely on the short RNA reads and requiring no genomic information is highly desirable. In this study, we propose an approach that relies primarily on the nucleotide composition of the read and does not require reference genomes of related species for prediction...

Over the past decade, high-throughput studies have identified many novel transcripts. While their existence is undisputed, their coding potential and functionality have remained controversial. Recent computational approaches guided by ribosome profiling have indicated that translation is far more pervasive than anticipated and takes place on many transcripts previously assumed to be non-coding. Some of these newly discovered translated transcripts encode short, functional proteins that had been missed in prior screens...

Lung cancer is a disease with a dismal prognosis and is the biggest cause of cancer deaths in many countries. Nonetheless, rapid technological developments in genome science promise more effective prevention and treatment strategies. Since the Human Genome Project, scientific advances have revolutionized the diagnosis and treatment of human cancers, including thoracic cancers. The latest, massively parallel, next generation sequencing (NGS) technologies offer much greater sequencing capacity than traditional, capillary-based Sanger sequencing...

BACKGROUND: Recent discoveries on bacterial transcriptomes gave evidence that small RNAs (sRNAs) have important regulatory roles in prokaryotic cells. Modern high-throughput sequencing approaches (RNA-Seq) enable the most detailed view on transcriptomes offering an unmatched comprehensiveness and single-base resolution. Whole transcriptome data obtained by RNA-Seq can be used to detect and characterize all transcript species, including small RNAs. Here, we describe an RNA-Seq approach for comprehensive detection and characterization of small RNAs from Corynebacterium glutamicum, an actinobacterium of high industrial relevance and model organism for medically important Corynebacterianeae, such as C...