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Affymetrix® Transcriptome Analysis Console Software

Comprehensive exploration of the transcriptome

Research has shown that the tens of thousands of human genes contain hundreds of thousands of exons, which produce hundreds of thousands of different transcript isoforms. These transcript isoforms are produced when the exons of a gene may be included within, or excluded from, the final, processed messenger RNA produced from that gene. The five basic modes of alternative splicing include:

  • Exon skipping
  • Mutually exclusive exons
  • Alternative 5' donor sites
  • Alternative 3' acceptor sites
  • Intron retention

Until now, measuring and analyzing these transcript isoforms has been nearly impossible due to technology limitations, sample input requirements, and lack of analysis capabilities/tools.

Affymetrix' next-generation GeneChip® Human Transcriptome Array 2.0 has been designed to eliminate these technology and analysis hurdles. This new array provides you with the ability to go beyond gene-level expression profiling by providing the coverage required to accurately detect transcript isoforms within a family of transcripts. The unparalleled coverage of this array provides clinical and translational researchers with the deepest insight into complex biology available. This high-resolution array design contains an unprecedented >7.3 million distinct probes covering coding and non-coding transcripts. 70% of the probes on this array cover exons for coding transcripts, and the remaining 30% of probes on the array cover exon-exon splice junctions and non-coding transcripts. The unparalleled coverage of this array provides the deepest insight into all coding and non-coding transcripts available.

To ensure uniform coverage of the transcriptome, GeneChip Human Transcriptome Array 2.0 was designed with approximately ten probes per exon and four probes per exon-exon splice junction. This coverage ensures that you will obtain complete, accurate, and reproducible data with every experiment.

In order to make the analysis of this vast amount genetic data seamless, the probes are all arranged into probe sets that translate and summarize your data in to gene-level, exon level, splice-junction probe sets. The CEL files generated by these arrays are compatible with Affymetrix Expression Console™ Software (version 1.3) to enable simple data analysis.

In order to make the vast amount of data generated useful, Affymetrix has developed Transcriptome Analysis Console (TAC) Software. This software package simplifies the analysis of your data and provides you with the capability to easily visualize expression changes at the gene and exon level as well as drill down into alternatively spliced exons. For the first time ever, GeneChip Human Transcriptome Array 2.0 coupled with Expression Console Software and Transcriptome Analysis Console Software offers researchers a complete solution from data to decision making. GeneChip Human Transcriptome Array 2.0 was designed to aid in human disease research and clinical translational medicine by providing the most comprehensive view of the transcriptome offered in a commercial microarray and by supporting analysis solutions that take you to biologically meaningful results in days rather than months.

Content profile

The human transcriptome has been well sequenced over the course of the past two decades and Affymetrix has used this information to formulate the most comprehensive set of gene models existing today. Currently most users of RNA-Seq have to choose a single public source such as RefSeq to map their data. By compiling content from a variety of public data sources an investigator doesn’t have to choose which transcriptome to map against decreasing your chances of missing genes. In summary:

  • 29% of the genes contain transcripts from ALL of the public data sources for the main content.
  • 28% of the genes contain transcripts from ONLY ONE of the public data sources.

The GeneChip Human Transcriptome Array design includes both coding and non-coding genes built on transcript mappings from hg19 human reference sequence (GRCh37). More than 400,000 full length transcripts were combined from available public data sources including RefSeq NCBI, UCSC Genes, Vertebrate Genome Annotation (Vega) database, GENCODE, Ensembl, and the Mammalian Gene Collection. Additional long non-coding content was combined from the UCSC genome browser, noncode.org, and the Broad Human Body Map*.

Data from each source was complied and curated to reduce redundancies and create concise gene models. A similar design strategy, employed in 2006, when compared to RNA-seq, has shown that over 99% of the genes detected across many tissues were represented on that 2006 array.**

Data sources used to design and annotate the array
RefSeq
Ensembl
UCSC Known Genes
UCSC lincRNA transcripts
Vertebrate Genome Annotation (Vega) database
MGC Mammalian Gene Collection (v10)
www.noncode.org
lncRNA db
Broad Institute, Human Body Map lincRNAs and TUCP (transcripts of uncertain coding potential) catalog

Array content Protein coding content Non-protein coding content
Genes (transcript clusters) 44,699 22,829
Transcripts 245,349 40,914
Exons 560,472 109,930
Exon clusters 296,058 82,444
Spice junctions (probe sets) 339,146 0

Controls
ERCC probe set1,2 63
Background probes Antigenomic set
Poly-A controls2 dap, lys, phe, thr
Hybridization controls bioB, bioc, bioD, creX

1 Probe sets interrogating external RNA controls present in the Ambion® ERCC RNA Spike-In Control Mixes, P/N 4456740 and 4456739.

2 This array contains probe sets for both ERCC and Poly-A spike in-controls. Sequence homology between the two control mixes will result in cross-hybridization of target to the control probes on the array. It is important to use only one control.


Specifications
Sensitivity ≥1:100,000 (≥1.5 pM)
Correlation coefficient (intra-lot) ≥0.99
Detectable fold change 2-fold for 1:100,000 vs. 1:50,000
Dynamic range ~3 logs
Total RNA input required 50–500 ng
Probe feature size 5 µm
Probe length 25-mer
Average probes per gene1 180
Probes per exon2 (median) 10
Probes per splice junction (median) 4
Interrogated strand3 Sense
Hybridization volume 200 µl
Fluidics script FS450_0001

1 Single and double exon genes were brought up to a minimum of 30 probes total per gene.

2 Single exon transcript clusters contain a median of 20 probes per exon.

3 The probes tiled on the array are designed in the anti-sense orientation, requiring sense-strand labeled targets to be hybridized to the array. By convention, the array is called ST array representing the necessity of using sense targets (the labeled samples to be hybridized to the array).

References

* Moran N., et al. Integrative annotation of human large intergenic noncoding RNAs reveals global properties and specific subclasses. Genes Development 25(18):1915–1927 (2011).

** Weihong Xu, et al. Human transcriptome array for high-throughput clinical studies, PNAS 108(9):3707-3712 (2011).

 

 

"For Research Use Only. Not for use in diagnostic procedures."

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