Arbitrary primed PCR fingerprinting of RNA applied to mapping differentially expressed genes.
Differential gene expression between various tissues and developmental stages or between cells in vitro under different growth conditions can be rapidly and efficiently compared using the RNA arbitrarily primed polymerase chain reaction (RAP) fingerprinting method (Welsh et al., 1992b; Liang and Pardee, 1992). In RAP, a primer of arbitrary sequence primes both first and second strand cDNA synthesis. The mixture of products is then PCR amplified and resolved electrophoretically, yielding highly reproducible fingerprints that are tissue-specific or growth condition-specific. Differences between fingerprints arise from differentially expressed genes, as verified by Northern blot analysis. RAP can be performed on the RNA samples using various DNA primers. Each two day experiment yields a sample of approximately twenty cDNA products per lane making the identification of differentially or developmentally regulated genes no longer rate limiting. Those PCR products representing genes that are regulated can be cloned from the gel and sequenced. Sequences can be compared to the DNA and protein sequence databases to identify homologs, motifs and members of gene families. The clones can be placed on the genetic map as Expression Tagged Sites (ETS, Adams et al., 1991a).
McClelland
M
M
California Institute of Biological Research, La Jolla 92037.
Chada
K
K
Welsh
J
J
Ralph
D
D
D000818:Animals; D001483:Base Sequence; D002874:Chromosome Mapping / Q000379:methods*; D016172:DNA Fingerprinting / Q000379:methods*; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D016045:Human Genome Project; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D012313:RNA / Q000235:genetics*
D017931:DNA Primers; D018076:DNA, Complementary; D012313:RNA