Molecular microbiology – recent tutorial references and resources

Basics

  1. DNA extraction – essential first step. Quantity of DNA obtained can be assayed spectrophotometrically.
  2. Reverse transcriptase step – for detection of RNA viruses
  3. Nucleic acid amplification –  know how PCR works! This PCR animation is easy to understand.  Many other methods now that are more complex and are isothermal – do not require cyclers to change the temperature of the reaction.
  4. Detection of product – most assays rely on closed tube ‘real-time’ detection of product. There are a variety of nucleic acid probe methods described.   Have a look here for a short video instruction.  Quantification becomes possible and is used widely.
  5. DNA sequencing – this is a basic technique these days (see below)

Excellent background materials from ICPMR, NSW ( circa 2009): 

Molecular-diagnosis-in-microbiology-ferguson-2016: overview presentation.

Important concepts

  • pre-analytical and assay factors that affect sensitivity and specificity
  • quality control (see above presentation)
  • amplification product quantification and its usefulness

N.B. visit your local molecular lab if there is one and examine and understand all of the assays in use – for commercial assays, obtain the product inserts to read.  

Modern methods

  • Next generation sequencing methods: basically a way of doing massive parallel sequencing of short multiple sequences in the same sample
  • GenXpert and similar methods : of great relevance for Nepal, PNG and elsewhere
  • Multiplex commercial molecular platforms – e.g. Biofire filmarray
  • Whole genome analyses

 

About mdjkf

Microbiologist and Infectious Diseases Physician
This entry was posted in Microbiology, Part 2 M Med Micro topics, PNG and tagged . Bookmark the permalink.

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