Essential Real-Time PCR

The Essential Principles and Applications of the Real-Time PCR course is part of the Essential Short Course Series™ and is held over a period of four days.

PCR has revolutionised the practice of science. This course is structured to expose the delegate to the advantages of using real-time PCR technology and chemistries whilst empowering the delegate with hands-on experience of real-time PCR applications. Validation of real-time PCR assays and specific quality assurance factors will be highlighted. This course will enable scientists to understand the principles of designing an efficient real-time PCR assay, interpreting real-time PCR data and troubleshooting real-time PCR assays.

This course is essential for scientists who want to use this quantitative technology in research or diagnostics

This Essential Real-Time PCR Short Course is an advanced course. No basic PCR theory or practical skills will be revisited during this course. We strongly recommend that delegates only register for this Essential Real-Time PCR Short Course if they have comprehensive and detailed knowledge and experience in PCR.

Delegates who do not have these skills are strongly advised to attend the Essential PCR Short Course to ensure that they are familiar with the detailed PCR theory and appropriate practical experience, as well as applications and advanced molecular biology calculations.

Course Programme

Day 1

Introduction to real-time PCR theory


  • Principles
  • Absorption and emission of fluorophores; Fluorescence resonance energy transfer (FRET)
  • DNA intercalating dyes
  • Probe based detection strategies

Discussion of Practical Sessions

Practical Session I:   Hands-on real-time PCR experience

  • Delegates set up their own SYBR Green real-time PCR reactions

Day 2

Real-time PCR reaction setup

Standard curves

  • Construction of a standard curve: Biostatistics principles; Linear regression

Theory of melt curve analysis

  • Standard melt curve analysis
  • High resolution melt curve analysis

Practical Session II:    Hands-on real-time PCR experience

  • Delegates set up their own SYBR Green real-time PCR reactions

Real-time PCR optimisation

Analysis of Practical Sessions I and II

  • Analysis of SYBR Green real-time PCR results: Melt curve analysis
  • Compare results from practical sessions I and II
  • Identify factors affecting success of real-time PCR reactions: Primer-dimers; GLP aspects

Amplification efficiency

  • Impact of efficiency of a PCR assay on the quantification results
  • Role of amplification efficiency in validation of assays

Primer and probe design, and synthesis

Day 3

Practical Session III:   Hands-on real-time PCR experience

  • Delegates set up their own probe based real-time PCR reactions

Reverse transcription real-time PCR

Practical Session IV:   Hands-on real-time PCR experience

  • Delegates set up their own SYBR Green real-time PCR reactions: Generating standard curve

Real-time PCR instrumentation

Appropriate controls for real-time PCR reactions and assays

Day 4

Troubleshooting of real-time PCR reactions

Data analysis of real-time PCR reactions

  • Step-for-step analysis
  • Absolute quantification
  • Relative quantification
  • Allelic discrimination
  • Plus/Minus assays

Analysis of Practical Sessions III

  • Analysis of probe based real-time PCR results
  • Determine genotypes of samples

Analysis of Practical Sessions IV

  • Analysis of SYBR green real-time PCR results
  • Determine concentrations of unknown samples via absolute quantification method

The sequence of this programme is preliminary and may change.

Comments of previous delegates on our Essential Real-Time PCR™ short courses:

“The course material and presenters were fabulous, I learnt so much and real-time PCR is much less daunting now, thank you.”

“This was a great course, the presenters really knew what they were talking about and it was a real good learning experience.”

“I thoroughly enjoyed this course, I found it extremely beneficial and it will most definitely have an impact on my whole understanding (+ working ability) on this subject. I am very impressed with the professional way in which DNAbiotec carries itself.”

“A great course worth every cent paid for (it). “

“The course is very comprehensive and helpful.”