Figure 2

Telomerase activity of MINO and tumor tissues. Telomerase activity by telomerase repeat amplification protocol (TRAP) assay. Representative ethidium stained polyacrylamide gel electrophoresis of the PCR products resulting from protein extract incubation with artificial telomere repeat template showing a bright 50 bp band with 6 bp increasing length ladder. At the bottom of the gel is a 36 bp internal PCR control, with PCR optimized to be semi-competitive with the extended telomere repeats produced by the telomerase protein derived from each sample. The protein samples are diluted in buffer and then added to the reaction mixture to provide a final reaction mixture concentration as listed (top of each lane). Matched MINO and tumor samples were tested. (a) Lines 4w4 and 4w11 are shown, with normal mouse mammary epithelium control (separated from the stroma by partial tissue dissociation and centrifugation) shown in the right three lanes. (b) Quantitation by comparing the band intensity with the internal control for multiple samples is shown, with standard deviation of the mean depicted for each bar. Line 11 tumor had the highest levels, at a mean of 500 (not shown), and all samples except for line B MINO were statistically significantly different from the normal control. There was a high level of variability between assay runs, depicted by the size of the error bars, but the qualitative data were clear, as shown in the gel (panel a). bp, base pairs; MINO, mammary intraepithelial neoplasia outgrowth; TRAP, telomerase repeat amplification protocol.