Primer3Plus is an enhanced web interface for the Primer3 primer design program, offering a more user-friendly alternative to the original CGI scripts. It provides an intuitive interface using modern web technologies and was developed in collaboration with molecular biologists and technicians. The interface focuses on the task at hand, hiding detailed settings until they are needed. It supports various sequence formats, including FASTA, and allows users to store settings and primer sequences locally. Primer3Plus also enables users to send selected primers to an order form, integrating with laboratory ordering systems. The open architecture allows easy integration with external software. The Perl source code is available under the GPL license from SourceForge, and the interface is available at http://www.bioinformatics.nl/primer3plus. Primer3Plus includes five distinct design tasks: Detection, Cloning, Sequencing, Primer Check, and Primer List. The Detection task is similar to the original Primer3 interface, while the Cloning task allows precise control over primer ends. The Sequencing task designs primers for (re-)sequencing, with configurable spacing and overlap. The Primer Check task provides information on specific primers, and the Primer List task returns all possible primers that meet current settings. Primer3Plus uses HTML and JavaScript for interactivity and has been tested on various browsers. It runs on both Linux and Windows platforms and can be integrated with other applications. Future work includes adding features to extract and display sequence information. Primer3Plus is now part of the Primer3 SourceForge project, facilitating further development and user feedback. Funding for this work was provided by Wageningen University and Research Centre.Primer3Plus is an enhanced web interface for the Primer3 primer design program, offering a more user-friendly alternative to the original CGI scripts. It provides an intuitive interface using modern web technologies and was developed in collaboration with molecular biologists and technicians. The interface focuses on the task at hand, hiding detailed settings until they are needed. It supports various sequence formats, including FASTA, and allows users to store settings and primer sequences locally. Primer3Plus also enables users to send selected primers to an order form, integrating with laboratory ordering systems. The open architecture allows easy integration with external software. The Perl source code is available under the GPL license from SourceForge, and the interface is available at http://www.bioinformatics.nl/primer3plus. Primer3Plus includes five distinct design tasks: Detection, Cloning, Sequencing, Primer Check, and Primer List. The Detection task is similar to the original Primer3 interface, while the Cloning task allows precise control over primer ends. The Sequencing task designs primers for (re-)sequencing, with configurable spacing and overlap. The Primer Check task provides information on specific primers, and the Primer List task returns all possible primers that meet current settings. Primer3Plus uses HTML and JavaScript for interactivity and has been tested on various browsers. It runs on both Linux and Windows platforms and can be integrated with other applications. Future work includes adding features to extract and display sequence information. Primer3Plus is now part of the Primer3 SourceForge project, facilitating further development and user feedback. Funding for this work was provided by Wageningen University and Research Centre.