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Parameters for Successful PCR Primer Design

  • Stephen A. BustinEmail author
  • Reinhold Mueller
  • Tania Nolan
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 2065)

Abstract

Primers are critical components of any PCR assay, as they are the main determinants of its specificity, sensitivity, and robustness. Despite the publication of numerous guidelines, the actual design of many published assays is often unsound: primers lack the claimed specificity, they may have to compete with secondary structures at their binding sites, primer dimer formation may affect the assay’s sensitivity or they may bind only within a narrow temperature range. This chapter provides simple guidance to avoid these most common issues.

Key words

MIQE Oligonucleotide primers qPCR Assay design 

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Stephen A. Bustin
    • 1
    Email author
  • Reinhold Mueller
    • 2
  • Tania Nolan
    • 3
  1. 1.Faculty of Health, Education, Medicine, and Social CareAnglia Ruskin UniversityChelmsfordUK
  2. 2.RM ConsultingSan DiegoUSA
  3. 3.Faculty of Medical and Human Sciences, Institute of Population HealthUniversity of ManchesterManchesterUK

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