title: A synthetic biology standard for Chinese Hamster Ovary cell genome monitoring and contaminant detection by polymerase chain reaction.
abstract: BACKGROUND
  Chinese Hamster Ovary (CHO) cells are the current industry standard for production of therapeutic monoclonal antibodies at commercial scales. Production optimisation in CHO cells hinges on analytical technologies such as the use of the polymerase chain reaction (PCR) to quantify genetic factors within the CHO genome and to detect the presence of contaminant organisms. PCR-based assays, whilst sensitive and accurate, are limited by (i) requiring lengthy sample preparation and (ii) a lack of standardisation.
  
  
  RESULTS
  In this study we directly assess for the first time the effect of CHO cellular material on quantitative PCR (qPCR) and end-point PCR (e-pPCR) when used to measure and detect copies of a CHO genomic locus and a mycoplasma sequence. We also perform the first head-to-head comparison of the performance of a conventional qPCR method to that of the novel linear regression of efficiency (LRE) method when used to perform absolute qPCR on CHO-derived material. LRE qPCR features the putatively universal 'CAL1' standard.
  
  
  CONCLUSIONS
  We find that sample preparation is required for accurate quantitation of a genomic target locus, but mycoplasma DNA sequences can be detected in the presence of high concentrations of CHO cellular material. The LRE qPCR method matches performance of a conventional qPCR approach and as such we invite the synthetic biology community to adopt CAL1 as a synthetic biology calibration standard for qPCR.
authors.affiliation: Department of Biochemical Engineering, University College London, Bernard Katz Building, London, WC1E 6BT UK.
authors.forename: Alexander
authors.initials: A
authors.lastname: Templar
authors.affiliation: Department of Biochemical Engineering, University College London, Bernard Katz Building, London, WC1E 6BT UK.
authors.forename: Douglas
authors.initials: D
authors.lastname: Marsh
authors.affiliation: Department of Biochemical Engineering, University College London, Bernard Katz Building, London, WC1E 6BT UK.
authors.forename: Darren N
authors.initials: DN
authors.lastname: Nesbeth
keywords: Mammalian cells; PCR; Standardisation; Synthetic biology