Fig. 10.
Comparison of SC qPCR and LRE qPCR methods for absolute quantitation of a mycoplasmal DNA sequence. SC qPCR (a) and LRE qPCR (b) methods were used to quantify the number of copies of a mycoplasmal DNA sequence present in reactions containing 5 ng plasmid DNA (1.54 × 109 copies) plus disrupted CHO cells derived from a 2 × 106 cells/mL shake flask sample or 2.5 × 105, 6 × 106 and 1 × 107 cells/mL bioreactor cultivation samples (see legend). The number of copies of the plasmid in a given sample was also inferred from spectrophotometric measurement of total DNA concentration before addition of disrupted cells. These spectrophotometric measurements (not indicated for clarity) are linearly regressed (thick dashed line) for qualitative comparison. For LRE qPCR (b) it is possible to assess quantification of the pure mycoplasmal sequence (open squares) because the unrelated CAL1 reaction is used for calibration. For SC qPCR (a), quantification of the pure mycoplasmal sequence is not informative as this reaction represents the standard curve used for calibration