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Advanced qPCR Techniques for Publication Success: Following MIQE Recommendation

 

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Overview

The real-time reverse transcription (RT) polymerase chain reaction (PCR) (RT-qPCR) and real time PCR methods address the evident requirement for quantitative data analysis in molecular medicine, biotechnology, microbiology, diagnostics and other areas and have become the methods of choice for the quantification of nucleic acid targets and identification of sequence specific variations. Although often described as a “gold” standard, these are far from being routine assays.


Lecture and Lab Topics

  • Introduction to MIQE
  • qPCR Assay Design and Optimization 

 

  • Effect of RNA Degradation on RT-qPCR
  • miRNA Degradation
  • Effect of Inhibitors on RT-qPCR
  • Standard RNA Assessments and Quantification
  • RNA Integrity (5'/3')
  • Sample Quality (SPUD)

 

  • Reference Gene Selection
  • Quantification Strategies
  • Normalization Approaches for microRNA
  • Using Normalization to Assess Sample Quality

 

  • Genomic Regulation of Gene Expression
  • qPCR Gene Expression Experiments
  • Lessons for the MMR Scandal
  • QC prior to Publication
  • Biostatistics
  • Data analysis
  • Troubleshooting
  • Interpretation of ZFN Data


Previous Guest Lecturers

Tania Nolan – Sigma-Aldrich, Haverhill, UK
Stephen Bustin – Centre for Academic Surgery, Institute of Cell and Molecular Science, Barts and London School of Medicine and Dentistry, London, UK
Jo Vandesompele – Centre for Medical Genetics, Ghent University Hospital, Ghent, Belgium
Michael Pfaffl – Physiology Weihenstephan, Technical University Munich, Freising, Germany
Mikael Kubista – TATAA Biocenter, Goteborg, Sweden / Institute of Biotechnology AS CR, Prague, Czech Republic
Jim Huggett – LGC Limited, London, UK
Vladimir Benes – Genomic Core Facility, EMBL Heidelberg, Heidelberg, Germany


Feedback from Attendees

“This was a really helpful course, nice people, good organization. I will spread word! Thank you.”

“This course gave me courage to do future qPCR experiments in concordance with the MIQE guidelines. Now I know how to use the MIQE guidelines in practical work.”

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