Ensembl 2002: accommodating comparative genomics

Clamp, Michele; Andrews, Dan; Barker, Darren; Bevan, A. Paul; Cameron, Graham; Chen, Y.; Clark, Louise; Cox, Tony; Cuff, James; Curwen, V.; Down, Thomas A.; Durbin, Richard; Eyras, Eduardo; Gilbert, James; Hammond, Martin; Hubbard, Tim; Kasprzyk, Arek; Keefe, Damian; Lehväslaiho, Heikki; Iyer, Vivek; Melsopp, Craig; Mongin, Emmanuel; Pettett, Roger; Potter, Simon; Rust, Alistair G.; Schmidt, Erwin R.; Searle, Stephen M. J.; Slater, Guy; Smith, James; Spooner, William; Stabenau, Arne; Stalker, Jim; Stupka, Elia; Ureta-Vidal, Abel; Västrik, Imre; Birney, Ewan

doi:10.1093/nar/gkg083

Cited by 225 publications

(152 citation statements)

References 14 publications

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“…siRNAs can potentially hybridize with multiple RNA target genes and produce misleading experimental results [39]. To identify potential cross-hybridization we run BLAST against cDNA databases such as Unigene [40] or Ensembl [41] without any filtering. As a guideline, 19 bp siRNA sequences with identical matches of length P16 to other genes should be discarded.…”

Section: Resultsmentioning

confidence: 99%

Improved and automated prediction of effective siRNA

Chalk

Wahlestedt

Sonnhammer

2004

Biochemical and Biophysical Research Communications

119

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Short interfering RNAs are used in functional genomics studies to knockdown a single gene in a reversible manner. The results of siRNA experiments are highly dependent on the choice of siRNA sequence. In order to evaluate siRNA design rules, we collected a database of 398 siRNAs of known efficacy from 92 genes. We used this database to evaluate previously proposed rules from smaller datasets, and to find a new set of rules that are optimal for the entire database. We also trained a regression tree with full crossvalidation. It was however difficult to obtain the same precision as methods previously tested on small datasets from one or two genes. We show that those methods are overfitting as they work poorly on independent validation datasets from multiple genes. Our new design rules can predict siRNAs with efficacy P50% in 91% of cases, and with efficacy P90% in 52% of cases, which is more than a twofold improvement over random selection. Software for designing siRNAs is available online via a web server at http:// sisearch.cgb.ki.se/ or as a standalone version for high-throughput applications. Ó 2004 Elsevier Inc. All rights reserved.RNAi is a recently discovered biological phenomenon whereby a single gene may be inhibited at the RNA stage of synthesis. short interfering RNAs (siRNAs) are duplexes of two RNA molecules, typically 21-mers with a 2nt 3 0 overhang [1]. One strand is loaded into the RISC complex [2] after which a sequence specific cleavage of the target takes place. The strength of the principle behind any form of RNA targeting (siRNA and antisense) is that the molecule can be used to inhibit expression of any mRNA, and thus the protein it encodes. This effect can be demonstrated without affecting related proteins, making it an invaluable tool for functional genomics. siRNAs have been found to be effective in Arabidopsis thaliana, Drosophila melanogaster, Caenorhabditis elegans, and mammals [3]. Many reviews of the biological processes behind siRNA inhibition exist, see for example [3,4].Efficient reliable design of high efficacy siRNA molecules is essential to meet the needs for cost-effective high-throughput functional genomics projects. To meet these needs the siRNAs designed should at least conform to the following criteria: (a) be predicted with high accuracy, (b) be sequence specific, and (c) be produced in a form that facilitates high-throughput production. In this paper we address these criteria, focusing primarily on a and b.Despite the apparent ease of designing siRNAs (compared to a popular DNA-based knockdown technique: antisense oligonucleotides (AOs)), a number of problems still remain. Randomly selected siRNAs produce knockdown P50% with 58-78% success rate, while very effective siRNAs ( P90/95%) are found by chance 11-18% of the time [5,6].Initial design paradigms for siRNAs were based on motif rules, such as AAN (19)

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Section: Resultsmentioning

confidence: 99%

Improved and automated prediction of effective siRNA

Chalk

Wahlestedt

Sonnhammer

2004

Biochemical and Biophysical Research Communications

119

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“…Putative promoter sequences corresponding to all known murine and human genes were extracted from the mouse and human genome sequence, respectively, using a Perl script based on the application programming interface developed by the Ensembl project (v13, May 2003 release) (Clamp et al, 2003). For each gene flagged as a 'known gene', a genomic sequence was extracted, spanning from 1000 bp upstream to 200 bp downstream of the gene's putative transcription start site (TSS).…”

Section: Computational Promoter Analysismentioning

confidence: 99%

Parallel induction of ATM-dependent pro- and antiapoptotic signals in response to ionizing radiation in murine lymphoid tissue

Rashi-Elkeles

Elkon

Weizman³

et al. 2005

Oncogene

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The ATM protein kinase, functionally missing in patients with the human genetic disorder ataxia-telangiectasia, is a master regulator of the cellular network induced by DNA double-strand breaks. The ATM gene is also frequently mutated in sporadic cancers of lymphoid origin. Here, we applied a functional genomics approach that combined gene expression profiling and computational promoter analysis to obtain global dissection of the transcriptional response to ionizing radiation in murine lymphoid tissue. Cluster analysis revealed a prominent pattern characterizing dozens of genes whose response to irradiation was Atm-dependent. Computational analysis identified significant enrichment of the binding site signatures of NF-jB and p53 among promoters of these genes, pointing to the major role of these two transcription factors in mediating the Atm-dependent transcriptional response in the irradiated lymphoid tissue. Examination of the response showed that pro-and antiapoptotic signals were simultaneously induced, with the proapoptotic pathway mediated by p53 targets, and the prosurvival pathway by NF-jB targets. These findings further elucidate the molecular network induced by IR, point to novel putative NF-jB targets, and suggest a mechanistic model for cellular balancing between pro-and antiapoptotic signals induced by IR in lymphoid tissues, which has implications for cancer management. The emerging model suggests that restoring the p53-mediated apoptotic arm while blocking the NF-jB-mediated prosurvival arm could effectively increase the radiosensitivity of lymphoid tumors.

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“…For the IgSF members, the whole sequences from amphioxus are used to search against the proteomes of the model organisms. Considering the proteome of ascidian is unavailable, TBLASTX program was used to search against the ascidian cDNA database, which is from ͗http://ghost.zool.lyoto-u.ac.jp/indexrl.html͘, and the ENSEMBL server was used to predict the homologous genes to refine our analyses (29).…”

Section: Bioinformatics Analysesmentioning

confidence: 99%

Genes “Waiting” for Recruitment by the Adaptive Immune System: The Insights from Amphioxus

Yu¹,

Dong²,

Wu³

et al. 2005

The Journal of Immunology

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In seeking evidence of the existence of adaptive immune system (AIS) in ancient chordate, cDNA clones of six libraries from a protochordate, the Chinese amphioxus, were sequenced. Although the key molecules such as TCR, MHC, Ig, and RAG in AIS have not been identified from our database, we demonstrated in this study the extensive molecular evidence for the presence of genes homologous to many genes that are involved in AIS directly or indirectly, including some of which may represent the putative precursors of vertebrate AIS-related genes. The comparative analyses of these genes in different model organisms revealed the different fates of these genes during evolution. Their gene expression pattern suggested that the primitive digestive system is the pivotal place of the origin and evolution of the AIS. Our studies support the general statement that AIS appears after the jawless/jawed vertebrate split. However our study further reveals the fact that AIS is in its twilight in amphioxus and the evolution of the molecules in amphioxus are waiting for recruitment by the emergence of AIS.

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Ensembl 2002: accommodating comparative genomics

Cited by 225 publications

References 14 publications

Improved and automated prediction of effective siRNA

Improved and automated prediction of effective siRNA

Parallel induction of ATM-dependent pro- and antiapoptotic signals in response to ionizing radiation in murine lymphoid tissue

Genes “Waiting” for Recruitment by the Adaptive Immune System: The Insights from Amphioxus

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