Rapid identification of differentially expressed RNA transcripts in apoptotic T lymphocytes.
The mRNA differential display technique has several advantages over conventional subtractive hybridization due to its simplicity and high sensitivity. However, the relatively high incidence of 'false positive' bands makes it more difficult to identify cDNAs derived from differential display, especially when subsequent large scale screening is required to isolate specific genes of interest. In this paper, we describe a simple, fast and efficient procedure which includes two-cycle PCR based probe labeling and direct sequencing of differential display products using 5' arbitrary and 3' oligo(dT) anchored primers for the rapid identification of differential display products. As demonstrated by identification of up-regulated RNA transcripts in apoptotic T lymphocytes, we showed that this modified technique greatly simplified the original procedure for the identification of differential display products and facilitated the application of differential display technique in the isolation and characterization of differentially expressed genes.
Zhang
J
J
Department of Cellular and Molecular Pathology, Toronto Hospital Research Institute, University of Toronto, Ontario, Canada.
Zhang
L
L
D000818:Animals; D017209:Apoptosis / Q000235:genetics*; D001483:Base Sequence; D006825:Hybridomas; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D012313:RNA / Q000235:genetics*; D013601:T-Lymphocytes / Q000473:pathology*; D014158:Transcription, Genetic
D012313:RNA