The quantification standard is genomic DNA extracted from low passage and defined mycoplasma or legionella cultivated in liquid medium according to the European Pharmacopoeia or in Hayflick"s medium. The extraction of the DNA is done by column absorption methods and the DNA purified by phenol/chloroform extraction and ethanol precipitation. The concentration was determined photometrically and by real-time PCR. The DNA concentration was adjusted by dilution with DNA stabilizing buffer. Genomic DNA is used as a control template when DNA amplification tests by conventional PCR are performed (e.g. as contamination control in cell culturing). With the real-time PCR technology, this titrated DNA standard with a defined concentration of genome copies is used as calibrator to generate standard curves. Therefore the provided solution is diluted in a 10x dilution series with DNA-free water and used as a PCR sample. With the generated fluorescent data, the software provided with the real-time PCR instrument calculates a standard curve, which can be used to determine the DNA concentration of unknown samples.

Contents

1 vial (green cap) of Standard DNA, 100 µl, contains approx. 106 genomes/µl.3 vials (white cap) with 1 ml of DNA free water.

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Protocols

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Dumke R. et al., (2017). Multi-center evaluation of one commercial and twelve in-house real-time PCR assays for detection of Mycoplasma pneumoniae. Diagnostic Microbiology & Infectious Disease, doi: 10.1016/j.diagmicrobio.2017.03.004.

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Applications

These DNA-Standards allow for the performance of a control reaction for PCR using qPCR instrumentation and to create a standard curve for quantification experiments, i.e. with Venor^®Mp Mycoplasma pneumoniae Diagnostic Kit for qPCR. The DNA standards can be used as a general control to confirm PCR system performance using a qPCR instrument or for the validation of detection limits in respect to conventional PCR tests.

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