Dataset Information

The chemiluminescence based Ziplex automated workstation focus array reproduces ovarian cancer Affymetrix GeneChip expression profiles.

ABSTRACT:

Background

As gene expression signatures may serve as biomarkers, there is a need to develop technologies based on mRNA expression patterns that are adaptable for translational research. Xceed Molecular has recently developed a Ziplex technology, that can assay for gene expression of a discrete number of genes as a focused array. The present study has evaluated the reproducibility of the Ziplex system as applied to ovarian cancer research of genes shown to exhibit distinct expression profiles initially assessed by Affymetrix GeneChip analyses.

Methods

The new chemiluminescence-based Ziplex gene expression array technology was evaluated for the expression of 93 genes selected based on their Affymetrix GeneChip profiles as applied to ovarian cancer research. Probe design was based on the Affymetrix target sequence that favors the 3' UTR of transcripts in order to maximize reproducibility across platforms. Gene expression analysis was performed using the Ziplex Automated Workstation. Statistical analyses were performed to evaluate reproducibility of both the magnitude of expression and differences between normal and tumor samples by correlation analyses, fold change differences and statistical significance testing.

Results

Expressions of 82 of 93 (88.2%) genes were highly correlated (p < 0.01) in a comparison of the two platforms. Overall, 75 of 93 (80.6%) genes exhibited consistent results in normal versus tumor tissue comparisons for both platforms (p < 0.001). The fold change differences were concordant for 87 of 93 (94%) genes, where there was agreement between the platforms regarding statistical significance for 71 (76%) of 87 genes. There was a strong agreement between the two platforms as shown by comparisons of log2 fold differences of gene expression between tumor versus normal samples (R = 0.93) and by Bland-Altman analysis, where greater than 90% of expression values fell within the 95% limits of agreement.

Conclusion

Overall concordance of gene expression patterns based on correlations, statistical significance between tumor and normal ovary data, and fold changes was consistent between the Ziplex and Affymetrix platforms. The reproducibility and ease-of-use of the technology suggests that the Ziplex array is a suitable platform for translational research.

SUBMITTER: Quinn MC

PROVIDER: S-EPMC2724495 | biostudies-literature | 2009 Jul

REPOSITORIES: biostudies-literature

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Publications

The chemiluminescence based Ziplex automated workstation focus array reproduces ovarian cancer Affymetrix GeneChip expression profiles.

Quinn Michael C J MC Wilson Daniel J DJ Young Fiona F Dempsey Adam A AA Arcand Suzanna L SL Birch Ashley H AH Wojnarowicz Paulina M PM Provencher Diane D Mes-Masson Anne-Marie AM Englert David D Tonin Patricia N PN

Journal of translational medicine 20090706

<h4>Background</h4>As gene expression signatures may serve as biomarkers, there is a need to develop technologies based on mRNA expression patterns that are adaptable for translational research. Xceed Molecular has recently developed a Ziplex technology, that can assay for gene expression of a discrete number of genes as a focused array. The present study has evaluated the reproducibility of the Ziplex system as applied to ovarian cancer research of genes shown to exhibit distinct expression pro ...[more]

PMID: 19580657

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Project description:BackgroundNatural antisense transcripts (NATs) are transcripts of the opposite DNA strand to the sense-strand either at the same locus (cis-encoded) or a different locus (trans-encoded). They can affect gene expression at multiple stages including transcription, RNA processing and transport, and translation. NATs give rise to sense-antisense transcript pairs and the number of these identified has escalated greatly with the availability of DNA sequencing resources and public databases. Traditionally, NATs were identified by the alignment of full-length cDNAs or expressed sequence tags to genome sequences, but an alternative method for large-scale detection of sense-antisense transcript pairs involves the use of microarrays. In this study we developed a novel protocol to assay sense- and antisense-strand transcription on the 55 K Affymetrix GeneChip Wheat Genome Array, which is a 3' in vitro transcription (3'IVT) expression array. We selected five different tissue types for assay to enable maximum discovery, and used the 'Chinese Spring' wheat genotype because most of the wheat GeneChip probe sequences were based on its genomic sequence. This study is the first report of using a 3'IVT expression array to discover the expression of natural sense-antisense transcript pairs, and may be considered as proof-of-concept.ResultsBy using alternative target preparation schemes, both the sense- and antisense-strand derived transcripts were labeled and hybridized to the Wheat GeneChip. Quality assurance verified that successful hybridization did occur in the antisense-strand assay. A stringent threshold for positive hybridization was applied, which resulted in the identification of 110 sense-antisense transcript pairs, as well as 80 potentially antisense-specific transcripts. Strand-specific RT-PCR validated the microarray observations, and showed that antisense transcription is likely to be tissue specific. For the annotated sense-antisense transcript pairs, analysis of the gene ontology terms showed a significant over-representation of transcripts involved in energy production. These included several representations of ATP synthase, photosystem proteins and RUBISCO, which indicated that photosynthesis is likely to be regulated by antisense transcripts.ConclusionThis study demonstrated the novel use of an adapted labeling protocol and a 3'IVT GeneChip array for large-scale identification of antisense transcription in wheat. The results show that antisense transcription is relatively abundant in wheat, and may affect the expression of valuable agronomic phenotypes. Future work should select potentially interesting transcript pairs for further functional characterization to determine biological activity.

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Dataset Information

The chemiluminescence based Ziplex automated workstation focus array reproduces ovarian cancer Affymetrix GeneChip expression profiles.

Background

Methods

Results

Conclusion

Publications

The chemiluminescence based Ziplex automated workstation focus array reproduces ovarian cancer Affymetrix GeneChip expression profiles.

Similar Datasets

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