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. 2009:2009:837170.
doi: 10.1155/2009/837170. Epub 2009 Oct 11.

Comparability of microarray data between amplified and non amplified RNA in colorectal carcinoma

Roland S Croner  1 , Berthold Lausen, Vera Schellerer, Isabel Zeittraeger, Axel Wein, Claus Schildberg, Thomas Papadopoulos, Arno Dimmler, Eckhart G Hahn, Werner Hohenberger, Wolfgang M Brueckl
Affiliations

Comparability of microarray data between amplified and non amplified RNA in colorectal carcinoma

Roland S Croner et al. J Biomed Biotechnol. 2009.

Abstract

Microarray analysis reaches increasing popularity during the investigation of prognostic gene clusters in oncology. The standardisation of technical procedures will be essential to compare various datasets produced by different research groups. In several projects the amount of available tissue is limited. In such cases the preamplification of RNA might be necessary prior to microarray hybridisation. To evaluate the comparability of microarray results generated either by amplified or non amplified RNA we isolated RNA from colorectal cancer samples (stage UICC IV) following tumour tissue enrichment by macroscopic manual dissection (CMD). One part of the RNA was directly labelled and hybridised to GeneChips (HG-U133A, Affymetrix), the other part of the RNA was amplified according to the "Eberwine" protocol and was then hybridised to the microarrays. During unsupervised hierarchical clustering the samples were divided in groups regarding the RNA pre-treatment and 5.726 differentially expressed genes were identified. Using independent microarray data of 31 amplified vs. 24 non amplified RNA samples from colon carcinomas (stage UICC III) in a set of 50 predictive genes we validated the amplification bias. In conclusion microarray data resulting from different pre-processing regarding RNA pre-amplification can not be compared within one analysis.

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Figures

Figure 1
Figure 1
Experimental procedure. RNA was isolated from tumours after CMD; one part of the RNA underwent amplification and the other part was hybridized without preamplification to microarrays.
Figure 2
Figure 2
Unsupervised (a) and supervised (b) hierarchical cluster analysis of microarray results from non amplified and amplified RNA samples from colorectal carcinoma samples.
Figure 3
Figure 3
Differences in total gene expressions between non amplified (NA) versus amplified (PA) RNA, regarding (a) sequence length of genes and (b) chromosomal localization.
Figure 4
Figure 4
Comparison of mean signals (HG-U 133A, Affymetrix) between RNA samples of 31 colon carcinomas amplified and 24 RNA samples of colon carcinomas not amplified prior to microarray hybridization.
See this image and copyright information in PMC

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