Modified Nucleotides in IVT: Small Changes, Big Impactย 

In our final blog post on double-stranded RNA (dsRNA), we turn our attention to the chemical building blocks of mRNA therapeuticsโ€”modified nucleotides. These seemingly minor changes to the RNA sequence play a crucial role in the success of mRNA-based vaccines and treatments. However, they also introduce complexities in accurately detecting and quantifying unwanted dsRNA byproductsโ€” key steps in ensuring the therapeutic efficacy of your mRNA product. 

What Are Modified Nucleotides? 

Modified nucleotides are ribonucleotides containing chemically altered nucleosides — like specialty ingredients swapped into a classic recipe to improve taste and nutrition. Just as a chef might use a lactose-free milk or gluten-free flour to make a dish easier to digest without changing its core structure, scientists use chemically altered nucleosides during in vitro transcription (IVT) to improve how mRNA therapies perform. These modifications replace their natural counterparts (e.g., uridine or cytidine) in the final RNA product. Their incorporation improves the performance and safety of mRNA therapeutics in several ways: 

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dsRNA QC Considerations: How I Learned to Stop Worrying and Love my IVT Reactions

As mRNA therapeutics continue to expand across clinical pipelines, one persistent challenge remains for developers: reducing double-stranded RNA (dsRNA) contaminants that can compromise safety and efficacy. These unintended byproducts of in vitro transcription (IVT) can trigger unwanted immune responses and reduce the potency of the final product. Developers must prioritize dsRNA detection and control as essential steps in the process. In our previous blog post we offered a high-level discussion of what is double-stranded RNA (dsRNA), its biological function, and importance of detection in a therapeutic context.  Here, weโ€™ll take a closer look at origins of dsRNA contamination, quality control measures, and improvement strategies.

Large-scale production of single-stranded RNA (ssRNA) for mRNA-based therapeutics is primarily done through in vitro transcription (IVT), an enzymatic process designed to generate high-yield, functional mRNA transcripts from a DNA template. This process uses purified RNA polymerase enzymes, such as T7, that recognize specific promoter sequences in the DNA template, generating the RNA transcripts of interest. However, IVT reactions also generate unwanted dsRNA byproduct. Below, we delve into some of the major quality control (QC) considerations and strategies to reduce dsRNA byproducts.

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Conjugate Like a Pro: Simplifying Antibody Labeling with On-Bead Conjugationย 

Antibody, On-bead conjugation

Labeled antibodies are indispensable tools in research and clinical diagnostics, used in everything from cell imaging and ELISAs to immunotherapies and ADC development. But if youโ€™ve ever tried labeling antibodies the traditional wayโ€”purify, buffer exchange, conjugate, purify againโ€”you know it can be tedious and time-consuming. Thatโ€™s where on-bead conjugation steps in with a solution. 

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Tuberculosis Genome Mapping in Italy: How 2,520 Strains Are Shaping the Future of TB Surveillance

Tuberculosis (TB) remains one of the deadliest infectious diseases globally, with millions of new cases and over a million deaths each year. The rise of drug-resistant strains has only complicated treatment and control efforts, turning TB into a moving target for clinicians and public health officials alike. Understanding how TB spreads, evolves and becomes resistant requires more than just microscopes and culturesโ€”it demands a detailed look at the bacteriumโ€™s genetic code.

Secondary tuberculosis in lungs and close-up view of Mycobacterium tuberculosis bacteria, 3D illustration
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Built for Whatโ€™s Next: Promega Expands Lyophilization to Meet Tomorrowโ€™s Demands

The new lyophilization equipment will more than double the lyophilization capacity of Promega Madison.

On March 12, 2025, a 46,000-pound stainless-steel chamber made a five-hour journey through Feynman Center to its final resting place in the brand-new Fill-Lyophilize-Finish suite. This massive piece of equipment will more than double the lyophilization capacity at Promega Madison, safeguarding the continuity of production and opening new frontiers in product formulation.

Lyophilization provides scientists with increased stability, enhanced flexibility and protection against error. Promega has been lyophilizing reagents in-house since the mid-1990s, and demand has steadily grown over time. The recent expansion reflects the companyโ€™s commitment to anticipating scientistsโ€™ future needs and planning for the long term.

Why is Lyophilization Important?

Lyophilization, also known as freeze-drying, provides a variety of benefits in the lab. For example, lyophilized reagents can typically be stored at higher temperatures, and they offer longer stability.

Stuart Forsyth inspects the lyophilization chamber during its installation.

โ€œLyophilized product also gives you added flexibility in how you tailor your reagents to your specific need,โ€ says Stuart Forsyth, Sr Process Validation Engineer at Promega. โ€œWhether youโ€™re reconstituting with a buffer, water or even a sample, youโ€™re able to alter the assayโ€™s concentration and formulation in ways that are impossible with liquid formulations.โ€

Many of the most popular Promega products include lyophilized components, including the CellTiter-Gloยฎ Luminescent Cell Viability Assay and ONE-Gloโ„ข Luciferase Assay System.

Promega also offers lyophilization for customers working with Promega to manufacture custom products. The flexibility helps many labs, especially diagnostics, ensure that the final reagent maximizes efficiency and ease of use for point-of-care applications.

โ€œEspecially if youโ€™re lyophilizing the whole assay in one, youโ€™re removing a lot of potential for mistakes by the user that would result in product failure,โ€ says Terri McDonnell, Director of Global Custom & OEM Commercial Development. โ€œLyophilization capabilities are powerful tools to have in your toolbox as you try to formulate a reagent for minimal risk of misuse or mistakes.โ€

Expanding Lyophilization at Promega Madison

The new lyophilizer will primarily be used with 10ml vials and 100ml bottles, but it can process numerous other formats.

Promega Operations closely monitors the throughput capacity of all critical processes. For years, the team has projected that manufacturing would outgrow the existing lyophilization capacity sometime in the mid-2020s. The project to build out the empty suite in Feynman Manufacturing Center began in 2021, and it will start producing products for sale in early 2026.

The new lyophilizer nearly doubles the throughput capacity of Promega Madison. It will primarily be used with 10ml vials and 100ml bottles, but the line can also handle 2ml and 3ml vials and large LyoGuard trays for bulk powder production. At this point, the team plans to primarily use the Feynman suite for high-demand catalog products like CellTiter-Glo, creating flexibility to use the older lines for custom products and other smaller demands.

Continuity, Collaboration and Creativity

The new lyophilization suite will have several significant impacts for scientists using Promega reagents.

First, the new lyophilization line creates additional redundancy to ensure that key products are continuously available. The huge increase in capacity means that if one lyophilizer is down for maintenance, the others can handle picking up the slack. The new suite also features the current state-of-the-art automation technology, minimizing any risks for contamination or human error that would disrupt high-quality production.

The lyophilizer is unloaded by crane outside Feynman Manufacturing Center.

For customers working with Promega on custom orders, the new lyophilizer gives Promega more flexibility to collaborate with customers on finding the right formulation for their needs, all within the established quality system.

“We partner with a wide range of customers seeking to adapt or customize our technologies for specific applications,โ€ says Terri McDonnell. โ€œAs the primary manufacturer of most of our products, and with the addition of new lyophilization capabilities, we can offer expanded scale and format options. Because these activities are performed in-house, we maintain greater control over quality and supply chain logistics, helping to ensure the consistent and reliable delivery of products.”

Finally, the additional capacity means that high-volume products can be manufactured less frequently by scaling up batch sizes. This frees up human resources to explore process improvements and dedicate more time to work outside of the production workflow. Kris Pearson, Director of Manufacturing Sciences and Custom operations, says the smaller equipment can serve as a sandbox where teams can test creative ideas.

โ€œWeโ€™ll have more opportunity to work with R&D on new product development, and to dive deep into new cycles and what that can mean for our custom capabilities,โ€ she says. โ€œWe can play around with new formats and processes to find new ways of offering a great product for every custom customer.โ€

Long-Term Planning and Strategy

As a private company, Promega isnโ€™t beholden to short-term gains. Leadership prioritizes decisions that support future needs, while building in room to adapt to changes in the scientific landscape.

The architectural drawings of Feynman Manufacturing Center show the suite earmarked for lyophilization as early as 2012, before the building was constructed.

โ€œWhen we started designing Feynman Manufacturing Center, we said we wanted 30% of the square footage to be frontier space,โ€ says Jen Romanin VP of Global Support and IVD Operations, and key member of the Global Planning Team. โ€œThis space would give us future flexibility in where new features would be installed.โ€

Sometimes needs are forecasted far in advance โ€“ for example, the architectural drawings of Feynman Manufacturing Center dated February 2012 show the new suite was already earmarked for Lyophilization almost a decade before the construction project began. Other spaces are left intentionally unlabeled as a nod to the unknown needs that will emerge over time. Whatever arises, the flexibility and foresight built into Promega facilities will position the team to respond quickly โ€“ and build a high-quality solution โ€“ without having to break new ground.

โ€œI think this says two things about us,โ€ says Chuck York, Vice President of Operations at Promega. โ€œFirst, it says weโ€™re pretty confident weโ€™re going to be here for a long time. Secondly, it says that no matter what happens between now and then, we want to make sure weโ€™re prepared.โ€


Using Dual-Luciferase Assays to Identify the Role of Non-Coding RNAs in Disease

In recent years, non-coding RNAsโ€”especially microRNAs (miRNAs) and long non-coding RNAs (lncRNAs)โ€”have emerged as powerful regulators of cellular behavior. These molecules modulate gene expression, often by targeting mRNAs for translational suppression or degradation. Two recent studiesโ€”one focused on osteoarthritis and the other on 5-Fu-resistant colorectal cancerโ€”illustrate how these non-coding, regulatory RNAs operate within disease-relevant signaling networks, providing new points for therapeutic intervention.

lncRNA, long noncoding RNA

Both studies use the pmirGLO Dual-Luciferase miRNA Target Expression Vector to evaluation predicted miRNA activity.  This dual-luciferase system offers a clean and quantifiable way to validate miRNAโ€“mRNA interactions using a simple bioluminescent readout. By cloning the 3ยด untranslated regions (UTRs) of suspected targets downstream of a firefly luciferase reporter and normalizing against Renilla luciferase, researchers can rapidly confirm whether a miRNA directly regulates its target.

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What Makes OBI-992 Different? A Closer Look at This TROP2 Antibody Drug Conjugate

ADC depiction

Antibody-drug conjugates (ADCs) are an increasingly powerful class of cancer therapeutics that combine the targeted precision of monoclonal antibodies with the cytotoxic potency of small-molecule drugs. By directing chemotherapy agents specifically to tumor cells, ADCs aim to maximize antitumor activity while minimizing damage to healthy tissues. One key challenge in ADC design is selecting the right target and payloadโ€”features that define efficacy, safety and resistance. 

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Mapping the Mind: In Vivo Imaging of Synaptic Plasticity with HaloTagยฎ Ligands

The brain is constantly rewiring itself, fine-tuning connections that shape how we think, learn, and remember. But capturing those fleeting molecular changes as they happen โ€” at the level of individual synapses and across entire brain regions โ€” has long been a challenge in neuroscience. Now, thanks to recent advances in HaloTagยฎ dye technology, researchers can visualize protein dynamics in living brains with stunning clarity and specificity.

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Bioluminescence vs. Fluorescence: Choosing the Right Assay for Your Experimentย 

From enzyme activity to gene expression, light-based assays have become foundational tools in life science research. Among these, fluorescence and bioluminescence are two of the most widely-used approaches for detecting and quantifying biological events. Both rely on the emission of light, but the mechanisms generating that lightโ€”and the practical implications for experimental designโ€”are quite different. 

Choosing between a fluorescence or bioluminescence assay isnโ€™t as simple as picking between two reagents off the shelf. Each has strengths and limitations depending on the application, instrumentation, and biological system. In this blog, weโ€™ll walk through how each method works, where they shine (and where they donโ€™t), and what to consider when deciding which approach is right for your experiment. 

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Immune Surveillance Meets Innovation: The Critical Need for dsRNA Detection

Todayโ€™s blog is written by guest blogger, Kai Hillman, Associate Product Marketing Manager at Promega.

RNA therapeutics have revolutionized modern medicine, offering groundbreaking solutions for diseases that were once deemed untreatable. These innovative treatments harness the power of RNA molecules to correct genetic anomalies and modulate protein expression, paving the way for personalized medicine. Among the many facets of RNA biology, double-stranded RNA (dsRNA) plays a pivotal role in cellular processes and immune surveillance.

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