Evaluation of Standard DNAs for Real-time PCR Quantification

Abstract

Adaptability of polymerase chain reaction (PCR) products to standard DNAs for real-time PCR quantification was evaluated by those standard curves. The threshold cycles (Ct values) of the same number of gene copies differed at -0.7-5.6 cycles between the genomic DNAs and the PCR products amplified from 16S rRNA genes of methanogens. This result indicates that the quantitative error occurs when PCR products are used as standard DNAs. Besides, the PCR efficiency (E) based on the slope of the standard curves was insufficient to evaluate the adaptability of PCR products as standard DNAs for real-time PCR quantification.