EasyPrimer: user-friendly tool for pan-PCR/HRM primers design. Development of an HRM protocol on wzi gene for fast Klebsiella pneumoniae typing: Scientific Reports

M. Perini, A. Piazza, S. Panelli, D. Di Carlo, M. Corbella, F. Gona, F. Vailati, P. Marone, D.M. Cirillo, C. Farina, G. Zuccotti, F. Comandatore

Research output: Contribution to journalArticlepeer-review

Abstract

In this work we present EasyPrimer, a user-friendly online tool developed to assist pan-PCR and High Resolution Melting (HRM) primer design. The tool finds the most suitable regions for primer design in a gene alignment and returns a clear graphical representation of their positions on the consensus sequence. EasyPrimer is particularly useful in difficult contexts, e.g. on gene alignments of hundreds of sequences and/or on highly variable genes. HRM analysis is an emerging method for fast and cost saving bacterial typing and an HRM scheme of six primer pairs on five Multi-Locus Sequence Type (MLST) genes is already available for Klebsiella pneumoniae. We validated the tool designing a scheme of two HRM primer pairs on the hypervariable gene wzi of Klebsiella pneumoniae and compared the two schemes. The wzi scheme resulted to have a discriminatory power comparable to the HRM MLST scheme, using only one third of primer pairs. Then we successfully used the wzi HRM primer scheme to reconstruct a Klebsiella pneumoniae nosocomial outbreak in few hours. The use of hypervariable genes reduces the number of HRM primer pairs required for bacterial typing allowing to perform cost saving, large-scale surveillance programs. © 2020, The Author(s).
Original languageEnglish
Article number1307
JournalSci. Rep.
Volume10
Issue number1
DOIs
Publication statusPublished - 2020

Keywords

  • article
  • consensus sequence
  • high resolution melting analysis
  • Klebsiella pneumoniae
  • nonhuman
  • polymerase chain reaction
  • bacterial genome
  • classification
  • genetics
  • genomics
  • Klebsiella infection
  • microbiology
  • phylogeny
  • procedures
  • software
  • outer membrane protein
  • primer DNA
  • Bacterial Outer Membrane Proteins
  • DNA Primers
  • Genome, Bacterial
  • Genomics
  • Klebsiella Infections
  • Phylogeny
  • Polymerase Chain Reaction
  • Software

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