Expression levels pre-T mRNAs in subsets of thymic precursors. Total RNA was isolated from sorted CD34⁺CD1a⁻, CD34⁺CD1a⁺, and CD4 ISP human thymic subpopulations (> 99% pure upon reanalysis) and analyzed for the levels of pT mRNA expression by semiquantitative RT-PCR as described in Materials and Methods (A). mRNA levels were compared with HPRT expression and calculated in arbitrary units. Standard curves were established by simultaneous amplification of 2-fold dilutions of cDNA prepared from RNA isolated from total thymus. PCR product of standard curves and samples were dot-blotted in duplicate onto nylon filters and hybridized with end-labeled oligo nucleotides shown on the right-hand site of the figure. (B) Measurement of the cpm/dot values was performed with a phosphoimager. Standard curves were calculated in which the first point of the curve, containing the highest amount of cDNA, was set arbitrarily to 1,000 U. Values of the samples were calculated in units. The ratios of (pT)/(HPRT) were determined and visualized in the bar graph.