Direct quantification of gene expression using capillary electrophoresis with laser-induced fluorescence.
Quantification of gene expression provides valuable information regarding the response of cells or tissue to stimuli and often is accomplished by monitoring the level of messenger RNA (mRNA) being transcribed for a particular protein. Although numerous methods are commonly used to monitor gene expression, including Northern blotting, real-time polymerase chain reaction, and RNase protection assay, each method has its own drawbacks and limitations. Capillary electrophoresis with laser-induced fluorescence (CE-LIF) can reduce protocol time, eliminate the need for radioactivity, and provide superior sensitivity and dynamic range for quantification of RNA. In addition, CE-LIF can be used to directly determine the amount of an RNA species present, something that is difficult and not normally accomplished using current methods. Gene expression is detected using a fluorescently labeled riboprobe specific for a given RNA species. This direct approach was validated by analyzing levels of 28S RNA and also used to determine the amount of discoidin domain receptor 2 mRNA in cardiac tissue.
Goldsmith
Jack G
JG
Department of Chemistry, University of South Carolina-Aiken, Aiken, SC 29801, USA.
Ntuen
Edidiong C
EC
Goldsmith
Edie C
EC
D001483:Base Sequence; D017931:DNA Primers; D019075:Electrophoresis, Capillary / Q000379:methods*; D015870:Gene Expression; D007834:Lasers; D009693:Nucleic Acid Hybridization; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D012339:RNA, Ribosomal, 28S / Q000032:analysis / Q000235:genetics; D013050:Spectrometry, Fluorescence / Q000379:methods*
D017931:DNA Primers; D012333:RNA, Messenger; D012339:RNA, Ribosomal, 28S