Organ- and tissue-specific expression of the low density lipoprotein receptor. Rapid quantitation by competitive...

A rapid and sensitive method of quantifying very low-abundant mRNA species by competitive reverse transcriptase-polymerase chain reaction (cRT-PCR) is presented. For each analysis, a defined amount of total cellular RNA is co-reverse transcribed and co-amplified with a titration series of in vitro synthesized RNA from a clone of the mRNA which carries an internal deletion. The equivalence point, which defines the number of specific mRNA molecules in the sample, can be determined using ethidium bromide stained gels of the reaction products. We present here three examples of the application of this new technique to the quantification of the low abundant mRNA for the low density lipoprotein (LDL) receptor. First, the regulation of LDL receptor mRNA expression by the HMG-CoA reductase inhibitor, pravastatin, has been analysed in vitro, in a human gastric tumour cell line. Second, LDL receptor mRNA from various bovine tissues has been quantified. Third, the expression of LDL receptor mRNA in human tissues originating from colorectal carcinoma and corresponding tumour-tree margins of surgical specimens has been measured.