Web-based design software streamlines multiplex PCR assay development

Despite being a fundamental necessity, multiplex PCR primer design can be an overwhelming process, with a vast array of available assay technologies, probe types and methodologies for researchers to choose from. This interview with Lindy McClelland, Senior Scientist at LGC Biosearch Technologies, explores the challenges researchers face during assay development and how LGC’s new tool is designed to help streamline this process.

When to choose a PCR design tool vs custom assay design service

"Decision fatigue is common during assay design,” McClelland explains. “There's many ways to approach it and there are many probe types and oligo modifications scientists may not be familiar with." Customers come to McClelland with not only feelings of overwhelm, but also misconceptions on how to choose the best solution for their work. "For example, some customers come in with the assumption that BHQplus™ is the best choice for them because ‘it's a plus’, not realizing that their particular application would actually benefit from our standard BHQ™ probes.” The different probe types accommodate the varying sensitivity and specificity needs of each assay. Some assays will benefit from short probes with elevated melting temperatures, while others will benefit from longer probes that can tolerate variations that may arise in mutation-prone targets.

This is where design tools come in, taking the guesswork out of probe type for a given application, and letting the software choose the optimal sequence of probes and primers for you. Customers can decide between self-service design tools and professional design services. Which type of tool you use is dependent on your specific needs and expertise levels.

For experienced researchers the path forward is clear. "If you're very familiar with designing assays, you know your target sequences and potential mutations or off-targets present in your testing paradigm, and you know what probe type is best suited for your assay goals, you're probably going to want that control and seek a design tool to create your oligo content," McClelland advises. "If you don't have that familiarity regarding the nature of your target, or you have additional challenges, then custom design services become invaluable.”

In fact, the foundation of successful assay design starts before using any software. "It may seem simple, but really understanding what you're trying to achieve and having a clear picture in mind from the start is critical," McClelland states. So, whether you are looking to scale up in the future, have the potential to move oligos to GMP manufacturing pedigree, or you’re seeking expert suggestions for very challenging targets using a custom assay design service coupled with a systematic assay optimization and validation approach can be invaluable for meeting performance expectations for years to come.

New free software for multiplex PCR assay design

To address these common challenges in assay design, LGC has recently launched PCR Forge, a custom design tool offering significant advancement in self-service assay design capabilities. McClelland’s enthusiasm about the new tool is palpable. "I'm very excited about PCR Forge,” she shares. “As someone who uses multiple assay design platforms when supporting customers, I've been piping in all of my tips and frustrations from other platforms into a new design tool, and I am really pleased with the outcome."

PCR Forge is designed with user experience in mind, eliminating tedious manual processes found in existing PCR tools, a fact that McClelland is keen to emphasize. "What's really cool is once the sequence is put into PCR Forge, you can interact directly with that sequence by highlighting within the text box to annotate instructions for your design.” This is a marked improvement to most existing tools which require you to manually count out blocks of DNA.

The platform supports multiple design types, handling PCR, qPCR with various probe types including BHQ, BHQnova™, and BHQplus, as well as genotyping applications. Users can input sequences through multiple methods: copy and paste, NCBI accession numbers, gene names, or RSID numbers for genotyping applications. PCR Forge also includes sophisticated data management features allowing researchers to save, organize and retrieve designs using customizable tags, eliminating the frustration of losing track of previous work in endless file lists.

The application possibilities for PCR Forge are limitless, with current customers looking at a range of targets, and the team at LGC is looking forward to many more collaborative developments with customers.

AI integration and automated validation in PCR design

Looking ahead, McClelland is excited about the potential of artificial intelligence. "AI technologies are just phenomenal. It has the potential to shave weeks off development time and help researchers determine what combinations will have the highest success rate.”

She sees a future with fully integrated workflows where researchers design assays with AI assistance and send validation plans to automated labs that execute optimization protocols and return comprehensive data packages, all without leaving their computers.

As assay design continues to evolve, the combination of intuitive software tools, expert services, and systematic approaches offers researchers multiple pathways to success. The development of advanced tools like PCR Forge will ensure researchers can efficiently carry out assay design.

Try PCR forge today and discover LGC's selection of probes, including a probe selection tool here.