SpliceCenter: a suite of web-based bioinformatic applications for evaluating the impact of alternative splicing on RT-PCR, RNAi, microarray, and peptide-based studies.
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.


CONCLUSIONS
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.
Ryan
Michael C
MC
Genomics & Bioinformatics Group, Laboratory of Molecular Pharmacology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA. mryan@tigerteamconsulting.com
Zeeberg
Barry R
BR
Caplen
Natasha J
NJ
Cleland
James A
JA
Kahn
Ari B
AB
Liu
Hongfang
H
Weinstein
John N
JN
D017398:Alternative Splicing; D035843:Biomedical Research / Q000379:methods*; D015342:DNA Probes / Q000145:classification; D003628:Database Management Systems; D030561:Databases, Nucleic Acid; D033181:Information Dissemination; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D010455:Peptides / Q000032:analysis / Q000737:chemistry; D034622:RNA Interference; D022821:RNA Splice Sites; D022661:RNA, Untranslated / Q000145:classification; D012107:Research Design; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D014158:Transcription, Genetic; D014584:User-Computer Interface
D015342:DNA Probes; D010455:Peptides; D022821:RNA Splice Sites; D022661:RNA, Untranslated