QUANTITATIVE-ANALYSIS OF GROWTH-HORMONE (GH) PRE-MESSENGER-RNA EXPRESSION IN CULTURED RAT ANTERIOR-PITUITARY-CELLS BY AN INTRON-SPECIFIC AND COMPETITIVE PCR METHOD

We have developed a novel method of quantifying growth hormone(GH) pre -mRNA expression in anterior pituitary cells. DNA-free total RNA extra cted from cultured rat anterior pituitary cells was reverse transcribe d(RT) to cDNA, and RT products were subsequently quantitated by compet itive PCR using intron-specific primers of rat GH gene. After 6-h of i ncubation in treated cells, dexamethasone(Dex) and triiodo-L-thyronine (T3) significantly increased GH pre-mRNA levels (3.2- and 2.2-fold com pared to nontreated cells, respectively). However, Northern blot analy sis did not detect significant changes in GH mRNA levels. After 24-h i ncubation with Dex and T3, significant increases in GH mRNA levels wer e detected on Northern blots, but GH pre-mRNA levels did not differ be tween treated and non-treated cells. These findings suggest that both Dex and T3 treatments rapidly increase GH pre-mRNA levels in normal so matotropes. This method has high sensitivity and widespread applicatio n to the analysis of pre-mRNAs of target genes.