Improving Alignment of Ambiguously Mapping Small RNAs
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ABSTRACT:
ORGANISM(S): Arabidopsis thaliana
PROVIDER: GSE76281 | GEO | 2016/05/25
SECONDARY ACCESSION(S): PRJNA306778
REPOSITORIES: GEO
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Project description:To determine the extent to which the major small RNA pathways functions across the Arabidopsis thaliana genome, small RNA populations from several tissues of wild-type (wt) and mutant plants were amplified by RT-PCR and sequenced using high-throughput 454 sequencing technology. Keywords: small RNAs, high-throughput sequencing
2007-02-01 | GSE6682 | GEO
Project description:To determine the extent to which the major small RNA pathways functions across the Arabidopsis thaliana genome, small RNA populations from several tissues of wild-type (wt) and mutant plants were amplified by RT-PCR and sequenced using high-throughput 454 sequencing technology. Keywords: small RNAs, high-throughput sequencing Amplicons were prepared by 5' and 3' adaptor ligation and RT-PCR using small RNA fractions from inflorescence tissue (containing stage 1-12 flowers) of wt Col-0 plants, mutants with defects in each DCL gene (dcl1-7, dcl2-1, dcl3-1, dcl4-2), and mutants with defects in each RDR gene for which a function has been established (rdr1-1, rdr2-1, rdr6-15). Amplicons from whole seedlings (3 day post-germinations) were prepared from Col-0 and rdr6-15 plants. Small RNA preparations from leaf samples of Col-O that were either uninoculated or inoculated by Pseudomonas syringae pv tomato (DC3000hrcC) for 1 hr and 3 hr were also sequenced.
2007-02-01 | E-GEOD-6682 | biostudies-arrayexpress
Project description:High-throughput sequencing of small RNAs (sRNA-seq) is a popular method used to discover and annotate microRNAs (miRNAs), endogenous short interfering RNAs (siRNAs), and Piwi-associated RNAs (piRNAs). One of the key steps in sRNA-seq data analysis is alignment to a reference genome. sRNA-seq libraries often have a high proportion of reads that align to multiple genomic locations, which makes determining their true origins difficult. Commonly used sRNA-seq alignment methods result in either very low precision (choosing an alignment at random), or sensitivity (ignoring multi-mapping reads). Here, we describe and test an sRNA-seq alignment strategy that uses local genomic context to guide decisions on proper placements of multi-mapped sRNA-seq reads. Tests using simulated sRNA-seq data demonstrated that this local-weighting method outperforms other alignment strategies using three different plant genomes. Experimental analyses with real sRNA-seq data also indicate superior performance of local-weighting methods for both plant miRNAs and heterochromatic siRNAs. The local-weighting methods we have developed are implemented as part of the sRNA-seq analysis program ShortStack, which is freely available under a general public license. Improved genome alignments of sRNA-seq data should increase the quality of downstream analyses and genome annotation efforts.
| S-EPMC4938663 | biostudies-literature
Project description:Rheumatic heart disease (RHD) remains a serious public health problem in developing countries. Atrial fibrillation (AF) is a medical complication of RHD. Although the understanding of disease pathogenesis has advanced in recent years, the key questions need to be addressed. Transfer RNA–derived small RNAs (tsRNAs) are a novel type of short non-coding RNAs that have potential regulatory functions in various physiological and pathological processes. The present study used tsRNAs sequencing to investigate the relationship between RHD and atrial fibrillation (AF). Three paired myocardial papilla were taken from six rheumatic RHD patients with AF (3 cases) or without AF (3 cases) from January 2016 to January 2017 in Xiangya Hospital, Central South University. A total of 219 precisely matched tsRNAs were identified, and 77 tsRNAs (fold change > 2.0 and P < 0.05) were differently changed. Three tsRNAs (AS-tDR-001269, AS-tDR-001363, AS-tDR-006049) were randomly selected and verified by qRT-PCR. The results of qRT-PCR were consistent with tsRNAs sequencing, suggesting the tsRNAs sequencing was reliable. Then bioinformatics analysis indicated that the target genes were abundant in regulation of transcription, DNA binding, intracellular. Most of the genes were predicted to interplay with cytokine-cytokine receptor by KEGG analysis. Our findings uncover the pathological process of AF in RHD through tsRNAs sequencing. This study provides new ideas for future research on elucidating the mechanism of AF in RHD and offers potential new candidates for the treatment and diagnosis of RHD.
2021-12-23 | GSE185581 | GEO
Project description:There are significant differences in the lncRNA expression profiles in Chinese patients with pulmonary adenocarcinoma. LncRNAs such as AK124939 may be anti-cancer factors related to the progress of pulmonary adenocarcinoma. RNA extracted from three paired pulmonary adenocarcinoma tissue and adjacent normal lung tissue specimens was used to synthesize double-stranded complementary DNA (cDNA) after labeling and hybridization. The cDNA was labeled and hybridized to the lncRNA expression microarray, and array data were analyzed for hierarchical clustering. Gene coexpression networks were constructed to identify interactions among genes. To validate the microarray findings, we measured the relative expression levels of four random differentially expressed lncRNAs in the same tissue used for microarray using real-time quantitative polymerase chain reaction (qRT-PCR). The expression level of one lncRNA AK124939, in the paired pulmonary adenocarcinoma/adjacent normal lung tissue of another 30 patients was measured using qRT-PCR. The experimental data were further analyzed and compared with clinical features.
2014-04-17 | E-GEOD-56850 | biostudies-arrayexpress
Project description:Background: Recent analyses implicate changes in the expression of the box C/D class of small nucleolar RNAs (snoRNAs) in several human diseases. Methods: Here we report the identification of potential novel RNA targets for box C/D snoRNAs in budding yeast, using the approach of UV crosslinking and sequencing of hybrids (CLASH) with the snoRNP proteins Nop1, Nop56 and Nop58. We also developed a bioinformatics approach to filter snoRNA-target interactions for bona fide methylation guide interactions. Results: We recovered 241,420 hybrids, out of which 190,597 were classed as reproducible, high energy hybrids. As expected, the majority of snoRNA interactions were with the ribosomal RNAs (rRNAs). Following filtering, 117,047 reproducible hybrids included 51 of the 55 reported rRNA methylation sites. The majority of interactions at methylation sites were predicted to guide methylation. However, competing, potentially regulatory, binding was also identified. In marked contrast, following CLASH performed with the RNA helicase Mtr4 only 7% of snoRNA-rRNA interactions recovered were predicted to guide methylation. We propose that Mtr4 functions in dissociating inappropriate snoRNA-target interactions. Numerous snoRNA-snoRNA interactions were recovered, indicating potential cross regulation. The snoRNAs snR4 and snR45 were recently implicated in site-directed rRNA acetylation, and hybrids were identified adjacent to the acetylation sites. We also identified 1,368 reproducible snoRNA-mRNA interactions, representing 448 sites of interaction involving 39 snoRNAs and 382 mRNAs. Depletion of the snoRNAs U3, U14 or snR4 each altered the levels of numerous mRNAs. Targets identified by CLASH were over-represented among these species, but causality has yet to be established. Conclusions: Systematic mapping of snoRNA-target binding provides a catalogue of high-confidence binding sites and indicates numerous potential regulatory interactions.
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Project description:Small molecule flexible alignment is a critical component of both ligand- and structure-based methods in computer-aided drug discovery. Despite its importance, the availability of high-quality flexible alignment software packages is limited. Here, we present BCL::MolAlign, a freely available property-based molecular alignment program. BCL::MolAlign accommodates ligand flexibility through a combination of pregenerated conformers and on-the-fly bond rotation. BCL::MolAlign converges on alignment poses by sampling the relative orientations of mutually matching atom pairs between molecules through Monte Carlo Metropolis sampling. Across six diverse ligand data sets, BCL::MolAlign flexible alignment outperforms MOE, ROCS, and FLEXS in recovering native ligand binding poses. Moreover, the BCL::MolAlign alignment score is more predictive of ligand activity than maximum common substructure similarity across 10 data sets. Finally, on a recently published benchmark set of 20 high quality congeneric ligand-protein complexes, BCL::MolAlign is able to recover a larger fraction of native binding poses than maximum common substructure-based alignment and RosettaLigand. BCL::MolAlign can be obtained as part of the Biology and Chemistry Library (BCL) software package freely with an academic license or can be accessed via Web server at http://meilerlab.org/index.php/servers/molalign .
| S-EPMC6598199 | biostudies-literature
Project description:The core components of RNA silencing effector complexes include small RNAs and members of the Argonaute (AGO) protein family. Arabidopsis encodes ten AGOs and complex population of small RNAs. It remains largely unknown how these small RNAs are recognized and recruited by each AGO complex. Here we purified four AGO complexes and identified small RNAs in each complex by deep sequencing. Keywords: Small RNA sorting Small RNAs were prepared from Arabidopsis AGO1, AGO2, AGO4 and AGO5 complexes, ligated to a 3' adaptor and a 5' acceptor sequentially, and then RT-PCR amplified. PCR products were reamplified using a pair of solexa cloning primers and then provided for sequencing. For technical details, see Qi, Y., He, X., Wang, X.J., Kohany, O., Jurka, J., and Hannon, G.J. 2006. Distinct catalytic and non-catalytic roles of ARGONAUTE4 in RNA-directed DNA methylation. Nature 443(7114): 1008-1012.
2008-04-05 | E-GEOD-10036 | biostudies-arrayexpress
Project description:Postoperative cognitive dysfunction (POCD) is one of the severe complications inducing low life quality and high mortality after surgery, especially in elderly patients.Here we probed differentially expressed circRNAs using microarray assay in POCD patients, aiming to find potential key circRNAs related to the occurrence of POCD. Subsequently, ten dysregulated circRNAs were confirmed via quantitative real-time polymerase chain reaction (qRT-PCR) in 10 paired samples. Then, Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed to probe the vital functions of dysregulated genes.
2020-07-14 | GSE147277 | GEO
Project description:The elucidation of the largely unknown transcriptome of small RNAs is crucial for the understanding of genome and cellular function. We report here the results of the analysis of small RNAs (< 50 nt) in the ENCODE regions of the human genome. Size-fractionated RNAs from four different cell lines (HepG2, HelaS3, GM06990, SK-N-SH) were mapped with the forward and reverse ENCODE high-density resolution tiling arrays. The top 1% of hybridization signals are termed SmRfrags (Small RNA fragments). Eight percent of SmRfrags overlap the GENCODE genes (CDS), given that the majority map to intergenic regions (34%), intronic regions (53%), and untranslated regions (UTRs) (5%). In addition, 9.6% and 16.8% of SmRfrags in the 5' UTR regions overlap significantly with His/Pol II/TAF250 binding sites and DNase I Hypersensitive sites, respectively (compared to the 5.3% and 9% expected). Interestingly, 17%-24% (depending on the cell line) of SmRfrags are sense-antisense strand pairs that show evidence of overlapping transcription. Only 3.4% and 7.2% of SmRfrags in intergenic regions overlap transcribed fragments (Txfrags) in HeLa and GM06990 cell lines, respectively. We hypothesized that a fraction of the identified SmRfrags corresponded to microRNAs. We tested by Northern blot a set of 15 high-likelihood predictions of microRNA candidates that overlap with smRfrags and validated three potential microRNAs ( approximately 20 nt length). Notably, most of the remaining candidates showed a larger hybridizing band ( approximately 100 nt) that could be a microRNA precursor. The small RNA transcriptome is emerging as an important and abundant component of the genome function.
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