SUMMARY: Advances in sequencing technology have greatly reduced the costs incurred in collecting raw sequencing data. Academic laboratories and researchers therefore now have access to very large datasets of genomic alterations but limited time and computational resources to analyse their potential biological importance. Here, we provide a web-based application, Cancer-Related Analysis of Variants Toolkit, designed with an easy-to-use interface to facilitate the high-throughput assessment and prioritization of genes and missense alterations important for cancer tumorigenesis. Cancer-Related Analysis of Variants Toolkit provides predictive scores for germline variants, somatic mutations and relative gene importance, as well as annotations from published literature and databases. Results are emailed to users as MS Excel spreadsheets and/or tab-separated text files. AVAILABILITY: http://www.cravat.us/

SUMMARY: SpliceSeq is a resource for RNA-Seq data that provides a clear view of alternative splicing and identifies potential functional changes that result from splice variation. It displays intuitive visualizations and prioritized lists of results that highlight splicing events and their biological consequences. SpliceSeq unambiguously aligns reads to gene splice graphs, facilitating accurate analysis of large, complex transcript variants that cannot be adequately represented in other formats. AVAILABILITY AND IMPLEMENTATION: SpliceSeq is freely available at http://bioinformatics.mdanderson.org/main/SpliceSeq:Overview. The application is a Java program that can be launched via a browser or installed locally. Local installation requires MySQL and Bowtie. CONTACT: [email protected] SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

SUMMARY: Thousands of cancer exomes are currently being sequenced, yielding millions of non-synonymous single nucleotide variants (SNVs) of possible relevance to disease etiology. Here, we provide a software toolkit to prioritize SNVs based on their predicted contribution to tumorigenesis. It includes a database of precomputed, predictive features covering all positions in the annotated human exome and can be used either stand-alone or as part of a larger variant discovery pipeline. AVAILABILITY AND IMPLEMENTATION: MySQL database, source code and binaries freely available for academic/government use at http://wiki.chasmsoftware.org, Source in Python and C++. Requires 32 or 64-bit Linux system (tested on Fedora Core 8,10,11 and Ubuntu 10), 2.5*≤ Python <3.0*, MySQL server >5.0, 60 GB available hard disk space (50 MB for software and data files, 40 GB for MySQL database dump when uncompressed), 2 GB of RAM.

MOTIVATION: Splice variation plays important roles in evolution and cancer. Different splice variants of a gene may be characteristic of particular cellular processes, subcellular locations or organs. Although several genomic projects have identified splice variants, there have been no large-scale computational studies of the relationship between number of splice variants and biological function. The Gene Ontology (GO) and tools for leveraging GO, such as GoMiner, now make such a study feasible. RESULTS: We partitioned genes into two groups: those with numbers of splice variants b (b=1,..., 10). Then we used GoMiner to determine whether any GO categories are enriched in genes with particular numbers of splice variants. Since there was no a priori 'appropriate' partition boundary, we studied those 'robust' categories whose enrichment did not depend on the selection of a particular partition boundary. Furthermore, because the distribution of splice variant number was a snapshot taken at a particular point in time, we confirmed that those observations were stable across successive builds of GenBank. A small number of categories were found for genes in the lower partitions. A larger number of categories were found for genes in the higher partitions. Those categories were largely associated with cell death and signal transduction. Apoptotic genes tended to have a large repertoire of splice variants, and genes with splice variants exhibited a distinctive 'apoptotic island' in clustered image maps (CIMs). AVAILABILITY: Supplementary tables and figures are available at URL http://discover.nci.nih.gov/OG/supplementaryMaterials.html. The Safari browser appears to perform better than Firefox for these particular items.

SUMMARY: LS-SNP/PDB is a new WWW resource for genome-wide annotation of human non-synonymous (amino acid changing) SNPs. It serves high-quality protein graphics rendered with UCSF Chimera molecular visualization software. The system is kept up-to-date by an automated, high-throughput build pipeline that systematically maps human nsSNPs onto Protein Data Bank structures and annotates several biologically relevant features. AVAILABILITY: LS-SNP/PDB is available at (http://ls-snp.icm.jhu.edu/ls-snp-pdb) and via links from protein data bank (PDB) biology and chemistry tabs, UCSC Genome Browser Gene Details and SNP Details pages and PharmGKB Gene Variants Downloads/Cross-References pages.

BACKGROUND: Over 60% of protein-coding genes in vertebrates express mRNAs that undergo alternative splicing. The resulting collection of transcript isoforms poses significant challenges for contemporary biological assays. For example, RT-PCR validation of gene expression microarray results may be unsuccessful if the two technologies target different splice variants. Effective use of sequence-based technologies requires knowledge of the specific splice variant(s) that are targeted. In addition, the critical roles of alternative splice forms in biological function and in disease suggest that assay results may be more informative if analyzed in the context of the targeted splice variant. RESULTS: A number of contemporary technologies are used for analyzing transcripts or proteins. To enable investigation of the impact of splice variation on the interpretation of data derived from those technologies, we have developed SpliceCenter. SpliceCenter is a suite of user-friendly, web-based applications that includes programs for analysis of RT-PCR primer/probe sets, effectors of RNAi, microarrays, and protein-targeting technologies. Both interactive and high-throughput implementations of the tools are provided. The interactive versions of SpliceCenter tools provide visualizations of a gene's alternative transcripts and probe target positions, enabling the user to identify which splice variants are or are not targeted. The high-throughput batch versions accept user query files and provide results in tabular form. When, for example, we used SpliceCenter's batch siRNA-Check to process the Cancer Genome Anatomy Project's large-scale shRNA library, we found that only 59% of the 50,766 shRNAs in the library target all known splice variants of the target gene, 32% target some but not all, and 9% do not target any currently annotated transcript. CONCLUSION: SpliceCenter http://discover.nci.nih.gov/splicecenter provides unique, user-friendly applications for assessing the impact of transcript variation on the design and interpretation of RT-PCR, RNAi, gene expression microarrays, antibody-based detection, and mass spectrometry proteomics. The tools are intended for use by bench biologists as well as bioinformaticists.

MOTIVATION: Affymetrix microarrays are widely used to measure global expression of mRNA transcripts. That technology is based on the concept of a probe set. Individual probes within a probe set were originally designated by Affymetrix to hybridize with the same unique mRNA transcript. Because of increasing accuracy in knowledge of genomic sequences, however, a substantial number of the manufacturer's original probe groupings and mappings are now known to be inaccurate and must be corrected. Otherwise, analysis and interpretation of an Affymetrix microarray experiment will be in error. RESULTS: AffyProbeMiner is a computationally efficient platform-independent tool that uses all RefSeq mature RNA protein coding transcripts and validated complete coding sequences in GenBank to (1) regroup the individual probes into consistent probe sets and (2) remap the probe sets to the correct sets of mRNA transcripts. The individual probes are grouped into probe sets that are 'transcript-consistent' in that they hybridize to the same mRNA transcript (or transcripts) and, therefore, measure the same entity (or entities). About 65.6% of the probe sets on the HG-U133A chip were affected by the remapping. Pre-computed regrouped and remapped probe sets for many Affymetrix microarrays are made freely available at the AffyProbeMiner web site. Alternatively, we provide a web service that enables the user to perform the remapping for any type of short-oligo commercial or custom array that has an Affymetrix-format Chip Definition File (CDF). Important features that differentiate AffyProbeMiner from other approaches are flexibility in the handling of splice variants, computational efficiency, extensibility, customizability and user-friendliness of the interface. AVAILABILITY: The web interface and software (GPL open source license), are publicly-accessible at http://discover.nci.nih.gov/affyprobeminer.

BACKGROUND: There are many fewer genes in the human genome than there are expressed transcripts. Alternative splicing is the reason. Alternatively spliced transcripts are often specific to tissue type, developmental stage, environmental condition, or disease state. Accurate analysis of microarray expression data and design of new arrays for alternative splicing require assessment of probes at the sequence and exon levels. DESCRIPTION: SpliceMiner is a web interface for querying Evidence Viewer Database (EVDB). EVDB is a comprehensive, non-redundant compendium of splice variant data for human genes. We constructed EVDB as a queryable implementation of the NCBI Evidence Viewer (EV). EVDB is based on data obtained from NCBI Entrez Gene and EV. The automated EVDB build process uses only complete coding sequences, which may or may not include partial or complete 5' and 3' UTRs, and filters redundant splice variants. Unlike EV, which supports only one-at-a-time queries, SpliceMiner supports high-throughput batch queries and provides results in an easily parsable format. SpliceMiner maps probes to splice variants, effectively delineating the variants identified by a probe. CONCLUSION: EVDB can be queried by gene symbol, genomic coordinates, or probe sequence via a user-friendly web-based tool we call SpliceMiner (http://discover.nci.nih.gov/spliceminer). The EVDB/SpliceMiner combination provides an interface with human splice variant information and, going beyond the very valuable NCBI Evidence Viewer, supports fluent, high-throughput analysis. Integration of EVDB information into microarray analysis and design pipelines has the potential to improve the analysis and bioinformatic interpretation of gene expression data, for both batch and interactive processing. For example, whenever a gene expression value is recognized as important or appears anomalous in a microarray experiment, the interactive mode of SpliceMiner can be used quickly and easily to check for possible splice variant issues.