Journal Article

MFEprimer: multiple factor evaluation of the specificity of PCR primers

Wubin Qu, Zhiyong Shen, Dongsheng Zhao, Yi Yang and Chenggang Zhang

in Bioinformatics

Volume 25, issue 2, pages 276-278
Published in print January 2009 | ISSN: 1367-4803
Published online November 2008 | e-ISSN: 1460-2059 | DOI:
MFEprimer: multiple factor evaluation of the specificity of PCR primers

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Summary: We developed a program named MFEprimer for evaluating the specificity of PCR primers based on multiple factors, including sequence similarity, stability at the 3 ′ -end of the primer, melting temperature, GC content and number of binding sites between the primer and DNA templates. MFEprimer can help the user to select more suitable primers before running either standard or multiplex PCR reactions. The cDNA and genomic DNA databases of 10 widely used species, as well as user custom databases, were used as DNA templates for analyzing primers specificity. Furthermore, we maintained a Primer3Plus server with a modified Primer3Manager for one-stop primer design and specificity checking.

Availability: The web service of MFEprimer is freely available at A stand-alone version of MFEprimer can be downloaded at Primer3Plus on our server is located at All programs are licensed under the General Public License (Free Software Foundation, 1991).


Supplementary information: Supplementary data are available at Bioinformatics online.

Journal Article.  1737 words.  Illustrated.

Subjects: Bioinformatics and Computational Biology

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