TRANSCRIPTOME 2002: From Functional Genomics to Systems
Poster PDF file
A Superparamagnetic Particle-Based Kit for the Purification of Fluorescent Dye-Labeled cDNA
Lisa R. Booth, Dwayne T. Campogan, Karin A. Hughes, Douglas A. Spicer, Amy L. Springer, Prolinx, Inc., Bothell, WA
Nucleic acid microarrays allow for the profiling of thousands of genes in a single experiment and are an increasingly valuable tool for analyzing gene expression. A typical microarray is composed of a 3 X 1” slide containing an array of nucleic acid fragments to which fluorescently-labeled cDNAs are hybridized. cDNA is the product of reverse transcription reactions where mRNA is converted to cDNA. The removal of excess dyes from these reverse transcription reactions is vital to the accuracy and efficiency of experimental results. Prolinx,Inc., has developed a new method using superparamagentic beads for the removal of excess Cy3- and Cy5-dye labeled dNTP precursors from these reactions. The protocol affords minimal loss of product, making it ideal when sample size is limited. High cDNA recoveries reduce the potential for introduction of variability, and concomitant loss of data reliability, by enabling replicates using the same cDNA sample and increase the possibility of improved detection of rare message. Studies presented will compare and contrast methods of cDNA purification in expression analysis microarrays. Effect of cDNA recovery on signal intensity and assay sensitivity will be included.
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