Differential display

Differential display (also referred to as DDRT-PCR or DD-PCR) is a laboratory technique that allows a researcher to compare and identify changes in gene expression at the mRNA level between two or more eukaryotic cell samples.[1] It was the most commonly used method to compare expression profiles of two eukaryotic cell samples in the 1990s.[1] By 2000, differential display was superseded by DNA microarray approaches.[2]

In differential display, first all the RNA in each sample is reverse transcribed using a set of 3′ "anchored primers" (having a short sequence of deoxy-thymidine nucleotides at the end) to create a cDNA library for each sample, followed by PCR amplification using arbitrary 3′ primers for cDNA strand amplification together with anchored 3′ primers for RNA strand amplification, identical to those used to create the library; about forty arbitrary primers is the optimal number to transcribe almost all of the mRNA. The resulting transcripts are then separated by electrophoresis and visualized, so that they can be compared.[1] The method was prone to error due to different mRNAs migrated into single bands, differences in less abundant mRNAs getting drowned by more abundant mRNAs,[1] sensitivity to small changes in cell culture conditions, and a tendency to amplify 3′ fragments rather than full mRNAs, and the necessity to use about 300 primers to catch all the mRNA.[3]:316–317 The method was first published in Science in 1992.[1][4]

References
  1. Peale FV, Jr; Gerritsen, ME (September 2001). "Gene profiling techniques and their application in angiogenesis and vascular development". The Journal of Pathology. 195 (1): 7–19. doi:10.1002/path.888. PMID 11568887.
  2. Pandey, A; Mann, M (15 June 2000). "Proteomics to study genes and genomes". Nature. 405 (6788): 837–46. doi:10.1038/35015709. PMID 10866210.
  3. Reece, Richard J. (2004). Analysis of genes and genomes. Chichester: Wiley. ISBN 978-0-470-84380-2.
  4. Liang, P; Pardee, AB (14 August 1992). "Differential display of eukaryotic messenger RNA by means of the polymerase chain reaction". Science. 257 (5072): 967–71. PMID 1354393.
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