Figure 1.
A direct correlation between the abundance of the 4.2 kb MnSOD transcript and percentage of S-phase cells. A. An illustration of the MnSOD transcript: CDS, coding sequence; PAS, polyadenylation signal; ARE: AU-rich sequence; arrows indicate primer sequence used in the quantitative PCR assay. B. Normal human fibroblasts (NHFs) were cultured for different days and harvested for flow cytometry analysis of the percentage of S-phase cells. Total cellular RNA isolated from duplicate dishes were analyzed for total MnSOD mRNA levels using a quantitative RT-PCR assay and primers representing the coding sequence. Fold change was calculated by first normalizing to 18S RNA levels and then relative to MnSOD mRNA levels in cells with 30 percent S-phase. Asterisks represent statistical significance compared to cells with 30 percent S-phase; n=3, p<0.05. C. Quantitative RT-PCR assay was repeated using primers designed to distinguish the mRNA levels of the two MnSOD transcripts (see materials and methods section).