Subtractive hybridization provides a means of depleting nucleotide sequences which are common to two mRNA populations, resulting in a relative enrichment of sequences unique to the experimental target tissue of interest. The success of any subtractive hybridization approach will be determined in large part by two characteristics of the mRNA population in the biological system to be investigated. First, the extent to which the abundance of a specific transcript differs between tissue types or within a tissue between experimental conditions. A second important parameter is the nucleotide sequence complexity of the mRNA populations in the tissues to be investigated. Since its introduction and early success in the identification of the T-cell receptor and other immune-related genes, the subtractive hybridization method has been a very productive technique for the isolation of messenger ribonucleic acids specific to tissue types and stages of development.
Since the nucleotide sequence of cholecystokinin (CCK) cDNA was found in the rat gene, we applied cDNA to quantitate the CCK mRNA. The size of the mRNA for a CCK precursor was 850 nucleotides in length using brain cytoplasmic RNA. There were no bands except CCK mRNA by Northern blot analysis. We also examined the developmental changes and regional distribution of CCK mRNA in rat brains by dot‐blot and gel‐blot hybridization using CCK cDNA as a probe. CCK mRNA was barely detectable in the fetal brain, but started to increase postnatally and attained the plateau level after 20–30 days. Further, the level of CCK mRNA was highest in the frontal cortex, followed by those of the hippocampus and striatum. The cerebellum contained only negligible CCK mRNA. These results are in agreement with those of CCK concentration in the corresponding brain areas and suggest a transcriptional control of CCK concentration.
