A protocol for differential display of mRNA expression using either fluorescent or radioactive labeling.
Since its invention in the early 1990s, differential display (DD) has become one of the most commonly used techniques for identifying differentially expressed genes at the mRNA level. Unlike other genomic approaches, such as DNA microarrays, DD systematically detects changes in mRNA profiles among multiple samples being compared without the need of any prior knowledge of genomic information of the living organism being studied. Here, we present an optimized DD protocol with a fluorescent digital readout as well as traditional radioactive labeling. The resulting streamlined fluorescent DD process offers an unprecedented accuracy, sensitivity and throughput in comprehensive and quantitative analysis of eukaryotic gene expression. Results usually can be obtained within days using a limited number of primer combinations, but a comprehensive DD screen may take weeks or months to accomplish, depending on gene coverage required and the number of differentially expressed genes present within a biological system being compared.
Liang
Peng
P
GenHunter Corporation, 624 Grassmere Park Dr, Suite 17, Nashville, Tennessee 37211, USA. peng.liang@vanderbilt.edu
Meade
Jonathan D
JD
Pardee
Arthur B
AB
D003001:Cloning, Molecular / Q000379:methods; D004587:Electrophoresis, Agar Gel / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D005453:Fluorescence; D020869:Gene Expression Profiling / Q000379:methods*; D012333:RNA, Messenger / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods
D012333:RNA, Messenger