title: Development of a quantitative real-time PCR assay for direct detection of growth of cellulose-degrading bacterium Clostridium thermocellum in lignocellulosic degradation.
abstract: OBJECTIVE
  Currently, there is no direct method for detecting Clostridium thermocellum in the insoluble medium. In this study, a quantitative real-time PCR assay was developed for the direct growth detection of C. thermocellum at the single-cell level in lignocellulosic biomasses.
  
  
  RESULTS
  The assay targeted the cipA gene and was able to distinguish C. thermocellum from other species with good reproducibility which quantitative detection limit was 10 cell equivalents (CE) per reaction. OD600-based counting and qPCR quantification of C. thermocellum cultured in soluble medium were compared and an excellent consistency was revealed, indicating the appropriateness of the developed qPCR method. Analysis based on yellow affinity substrate and fermentation products may incorrectly estimate its population.
  
  
  CONCLUSIONS
  The developed assay can serve as a specific, sensitive and reproducible method for the detection of C. thermocellum in lignocellulosic biomass at the single-cell level.
  
  
  CONCLUSIONS
  With the ability to rapidly detect C. thermocellum, this method will contribute substantially to the understanding of the lignocellulosic biomass degradation mechanism. Moreover, it can also be applied to detect C. thermocellum growth in certain co-culture system for the understanding of the metabolic interactions.
authors: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, China.
authors: H
authors: H
authors: Tang
authors: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, China.
authors: J F
authors: JF
authors: Ou
authors: School of Bioscience and Bioengineering, South China University of Technology, Guangzhou, China.|State Key Laboratory of Pulp and Paper Engineering, South China University of Technology, Guangzhou, China.|Guangdong Provincial Key Laboratory of Fermentation and Enzyme Engineering, South China University of Technology, Guangzhou, China.
authors: M J
authors: MJ
authors: Zhu
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics; D018533:Biomass; D048013:Acetivibrio thermocellus / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D005285:Fermentation; D008031:Lignin / Q000378:metabolism*; D008969:Molecular Sequence Data; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results
chemical_list: D001426:Bacterial Proteins; C036909:lignocellulose; D008031:Lignin
keywords: Clostridium thermocellum; cipA gene; detection; lignocellulosic biomass; quantitative real-time PCR