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Related Concept Videos

Real Time RT-PCR02:57

Real Time RT-PCR

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Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...
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A Duplex Digital PCR Assay for Simultaneous Quantification of the Enterococcus spp. and the Human Fecal-associated HF183 Marker in Waters
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Human fecal source identification with real-time quantitative PCR.

Orin C Shanks1, Lindsay Peed, Mano Sivaganesan

  • 1National Risk Management Research Laboratory, Office of Research and Development, U.S. Environmental Protection Agency, Cincinnati, OH, USA.

Methods in Molecular Biology (Clifton, N.J.)
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A new real-time quantitative PCR (qPCR) method detects human fecal pollution in water. This method targets a genetic marker from Bacteroides dorei, aiding in public health risk assessment for waterborne diseases.

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Area of Science:

  • Environmental microbiology
  • Public health
  • Molecular biology

Background:

  • Waterborne diseases pose a global health threat.
  • Human fecal contamination is a primary source of waterborne pathogens.
  • Accurate detection of fecal pollution is crucial for water safety.

Purpose of the Study:

  • To develop a real-time quantitative PCR (qPCR) method for identifying human fecal pollution in water.
  • To target a specific genetic marker of human-associated Bacteroides dorei for improved detection accuracy.

Main Methods:

  • Water sample collection and filtration.
  • DNA isolation with sample processing controls.
  • Real-time quantitative PCR (qPCR) amplification with internal amplification controls.
  • Quality control data analysis.

Main Results:

  • The study describes a validated qPCR protocol for detecting human fecal indicators.
  • The method successfully targets the Bacteroides dorei genetic marker.
  • Robust controls ensure reliable identification of human fecal pollution.

Conclusions:

  • This qPCR method provides a reliable tool for identifying human fecal contamination in ambient water.
  • The approach enhances the assessment of public health risks associated with water quality.
  • Effective monitoring of water sources can be improved using this molecular technique.