Addressing PCR Biases in Environmental Microbiology Studies

  • Rita Sipos
  • Anna Székely
  • Sára Révész
  • Károly Márialigeti
Part of the Methods in Molecular Biology book series (MIMB, volume 599)

Abstract

Each step of a molecular environmental microbiology study is prone to errors, though the qualitative and quantitative biases of PCR amplification could result in the most serious biases. One has to be aware of this fact, and well-characterized PCR biases have to be avoided by using target-optimized PCR protocols. The most important tasks are primer and thermal profile optimization. We have shown that primer mismatches, even in the case of universal primers, can cause almost complete missing of common taxa from clone libraries, for example. Similarly high annealing temperatures can drastically distort community composition of the sample in the PCR product. Strategies of primer selection and PCR thermal profile design are discussed in detail.

Key words

PCR bias environmental microbiology PCR optimization PCR primer selection PCR thermal protocol 

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Copyright information

© Humana Press 2010

Authors and Affiliations

  • Rita Sipos
    • 1
  • Anna Székely
    • 1
  • Sára Révész
    • 1
  • Károly Márialigeti
    • 1
  1. 1.Department of MicrobiologyEötvös Loránd UniversityBudapestHungary

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