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BMC Research Notes

, 1:21

First Online: 05 June 2008Received: 26 February 2008Accepted: 05 June 2008DOI: 10.1186-1756-0500-1-21

Cite this article as: Mengual, L., Burset, M., Marín-Aguilera, M. et al. BMC Res Notes 2008 1: 21. doi:10.1186-1756-0500-1-21

Abstract

BackgroundAn accurate gene expression quantification using TaqMan Arrays TA could be limited by the low RNA quantity obtained from some clinical samples. The novel cDNA preamplification system, the TaqMan PreAmp Master Mix kit TPAMMK, enables a multiplex preamplification of cDNA targets and therefore, could provide a sufficient amount of specific amplicons for their posterior analysis on TA.

FindingsA multiplex preamplification of 47 genes was performed in 22 samples prior to their analysis by TA, and relative gene expression levels of non-preamplified NPA and preamplified PA samples were compared. Overall, the mean cycle threshold CT decrement in the PA genes was 3.85 ranging from 2.07 to 5.01. A high correlation r between the gene expression measurements of NPA and PA samples was found mean r = 0.970, ranging from 0.937 to 0.994; p < 0.001 in all selected cases. High correlation coefficients between NPA and PA samples were also obtained in the analysis of genes from degraded RNA samples and-or low abundance expressed genes.

ConclusionWe demonstrate that cDNA preamplification using the TPAMMK before TA analysis is a reliable approach to simultaneously measure gene expression of multiple targets in a single sample. Moreover, this procedure was validated in genes from degraded RNA samples and low abundance expressed genes. This combined methodology could have wide applications in clinical research, where scarce amounts of degraded RNA are usually obtained and several genes need to be quantified in each sample.

Electronic supplementary materialThe online version of this article doi:10.1186-1756-0500-1-21 contains supplementary material, which is available to authorized users.

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Autor: Lourdes Mengual - Moisès Burset - Mercedes Marín-Aguilera - María José Ribal - Antonio Alcaraz

Fuente: https://link.springer.com/







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