Multiplex PCR Primer Design for Simultaneous Detection of Multiple Pathogens

  • Wenchao YanEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1275)


Multiplex PCR provides a powerful tool for simultaneous detection and discrimination of multiple pathogens or different subtypes of a causative agent from humans, animals, and plants in a single reaction, and saves time and cost in the clinical diagnostic laboratory. Here, we describe the specific protocol of multiplex PCR primer design for simultaneous identification of more than one target from a same specimen. Different sizes of amplicons and similar T m values of primer sets are essential to successfully develop a feasible multiplex PCR assay.

Key words

Multiplex PCR Primer design Simultaneous detection Multiple pathogens Different subtypes of one infectious agent Oligo primer analysis software Primer-BLAST tool Sizes of amplicons Tm values of primer sets 



This work was supported by National Foundation of Natural and Science (31001058) and the Doctor Startup Foundation of Henan University of Science and Technology (09001350).


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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Animal Quarantine Laboratory, College of Animal Science and TechnologyHenan University of Science and TechnologyLuoyangChina

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