Peptide nucleic acids (PNAs) represent a powerful class of synthetic DNA/RNA mimics where the natural, negatively-charged sugar-phosphate backbone is expertly replaced by a unique, electrically neutral N-(2-aminoethyl)glycine (pseudo-peptide) backbone. This transformative structural alteration is the key to PNA's superior performance, eliminating the electrostatic repulsion inherent to natural nucleic acid hybridization and enabling remarkably high binding affinity and increased thermal stability to complementary DNA or RNA targets. At CD BioGlyco, our PNA synthesis provides researchers and pharmaceutical developers with high-purity, custom PNA oligomers, harnessing these unique properties for critical applications in diagnostics, therapeutics, and fundamental research.
PNA synthesis is inherently challenging but is optimally executed via the solid-phase peptide synthesis (SPPS) methodology, which our platforms are designed to fully support. We utilize highly optimized and robust Fmoc/Boc-protected PNA monomers for sequential oligomer construction, a chemistry that facilitates efficient, high-fidelity chain elongation and minimizes byproduct formation. This foundational technology ensures we can routinely handle sequences of varying complexity and length.
Our commitment to efficiency extends to advanced reaction kinetics. We employ optimized coupling reagents and synthesis protocols, such as those demonstrating superior performance (e.g., PyOxim) over conventional agents, resulting in minimized truncation sequences and incomplete couplings, particularly for complex sequences. Furthermore, we incorporate heat-assisted SPPS methodologies during the critical coupling stages, a technique known to overcome steric hindrance and aggregation issues, thereby significantly enhancing overall coupling efficiency and ensuring the highest possible yields of full-length product.
The true value of PNA lies in its customizability. We are experts in synthesizing complex PNA conjugates, crucial for enhancing cellular uptake and bioavailability for in vivo applications. This includes the incorporation of cell-penetrating peptides (CPPs), lipophilic molecules, fluorescent dyes, and specialized ligands like GalNAc, positioning the PNA for targeted delivery in therapeutic research. We also offer chiral PNA analogues to fine-tune binding properties and enhance the detection of point mutations in diagnostic assays.
We provide a flexible and comprehensive PNA synthesis offering designed to meet the demands of early discovery through preclinical development, ensuring scalability and modification versatility for any project.
Routine synthesis scales range from research-grade quantities up to bulk-scale synthesis for larger in vivo and therapeutic research projects.
We offer customizable purity options, including standard purification and high-purity options, ensuring the correct grade for applications ranging from PCR clamping to clinical diagnostics research.
Our automated platform and advanced coupling chemistry are optimized for sequences up to 20 bases, while we maintain the capability to synthesize longer, more complex oligomers upon consultation, mitigating the decreasing synthesis efficiency typically associated with greater length.
We specialize in attaching a vast array of functional moieties, including but not limited to: fluorescent dyes (e.g., FAM, Cy3, Cy5), biotin, CPPs, lipophilic groups (e.g., cholesterol), and targeting ligands like GalNAc, critical for liver-targeted delivery.
Our core expertise in both peptide and PNA chemistry allows for the seamless construction of PNA-peptide hybrids, which are vital for enhancing solubility, biological stability, and targeted delivery applications.
Our expert team engages in detailed discussions with the client to evaluate the target sequence, desired scale, required modifications, and application goals. We provide specialized support for sequence design, including recommendations for length (e.g., typically 13–18 bases for high specificity) and modification placement to ensure maximum efficacy and success rate for the specific research aim.
The PNA chain is synthesized step-wise using automated, state-of-the-art SPPS synthesizers. Each coupling and deprotection cycle is meticulously monitored, often utilizing enhanced, temperature-controlled protocols to ensure superior yield and minimize unwanted side reactions.
The newly synthesized PNA oligomer is cleaved from the solid support resin, and the nucleobase protecting groups are removed. This critical step is performed under optimized chemical conditions to preserve the integrity of the PNA backbone and any incorporated labile modifications.
The crude PNA product undergoes multiple stages of preparative high-performance liquid chromatography (HPLC) to isolate the desired full-length oligomer from any truncated sequences or byproducts. We offer various purity levels, typically greater than 70% or 90% UV-purity, based on the client's specific application requirements.
Every PNA product is subjected to stringent quality assurance checks. Characterization includes analytical HPLC to confirm purity and mass spectrometry (typically MALDI-TOF MS) to verify the exact molecular weight and sequence.
Clients receive the final lyophilized product along with a comprehensive certificate of analysis (CoA), including the raw analytical data (HPLC and MS traces), batch-specific synthesis details, and recommended storage conditions.
Journal: Beilstein journal of organic chemistry
DOI: 10.3762/bjoc.17.116
Published: 2023
Results: This review provides a comprehensive 30-year retrospective on PNA, a synthetic DNA mimic with a neutral pseudopeptide backbone. The authors detail PNA's evolution from its initial design aimed at improving triplex-forming oligonucleotides to its current status as a versatile tool in research and diagnostics. The discussion covers the fundamental binding modes and structural features of PNA, highlighting its high affinity and sequence specificity for DNA and RNA. A significant portion is devoted to the extensive chemical modifications explored to enhance PNA's properties, such as solubility, binding affinity, and cellular delivery, including constrained backbones, novel nucleobases, and various conjugates like cell-penetrating peptides. The article critically assesses PNA's successful applications in molecular assays, such as PCR clamping and fluorescence in situ hybridization (FISH), contrasting them with the slower progress in therapeutic development. The central theme that emerges is that despite PNA's remarkable in vitro performance, its transition to clinical applications is still hindered by challenges in bioavailability, tissue-specific delivery, and endosomal entrapment, awaiting innovative chemical and biological solutions to unlock its full biomedical potential.
Molecular Diagnostics and Probes
High-affinity PNA probes are utilized in techniques like fluorescent in situ hybridization (FISH) and microarrays to achieve superior sensitivity and specificity for detecting target DNA or RNA sequences. The ability to use shorter probes with less background noise makes PNA-FISH ideal for identifying infectious pathogens and chromosomal anomalies.
Antisense and Antigenic Therapeutics
PNAs are powerful agents for modulating gene expression, capable of binding to complementary mRNA (antisense) to inhibit translation or invading dsDNA (antigene) to block transcription. Custom-synthesized PNAs can be designed to target specific genes implicated in cancer and genetic disorders, offering precise therapeutic potential.
miRNA Inhibition and Regulation
Due to PNA's strong binding affinity for complementary RNA, our custom PNA oligomers serve as highly effective inhibitors of microRNAs (miRNAs). This application is crucial for studying gene function and developing novel therapies that target aberrantly expressed tumor-suppressive or oncogenic miRNAs.
PCR and qPCR Clamping
PNAs are utilized as sequence-specific PCR blockers or clamps, where they bind to a specific sequence in the DNA template, inhibiting the amplification of a competitor sequence or wild-type allele. This allows for the highly sensitive detection and quantification of rare mutations or single-nucleotide polymorphisms (SNPs) amidst a vast excess of non-mutant DNA.
Unrivaled Sequence Specificity and Binding Affinity
Our PNAs exhibit superior binding to complementary DNA or RNA compared to conventional oligonucleotides due to the neutral backbone, making single-mismatch detection reliable for accurate diagnostics. This heightened specificity minimizes off-target effects in therapeutic and diagnostic assays, a critical feature.
Exceptional Enzymatic Resistance and Stability
The synthetic peptide-like backbone is entirely resistant to degradation by endogenous nucleases and proteases, granting our PNA products superior half-life and stability for both in vitro and challenging in vivo applications. This biological resilience is a key differentiator for successful therapeutic candidates.
Proprietary High-Yield Synthesis Protocols
By employing advanced chemical strategies, including the optimization of coupling reagents and temperature-shift protocols, we consistently achieve higher coupling efficiencies and overall yields, reducing the cost and time required for your downstream application.
Rigorous Multi-Stage Quality Control
We go beyond standard checks by providing comprehensive documentation, including detailed HPLC purity traces and precise mass spectrometry verification for every order, ensuring maximum confidence in the structural identity and quality of your synthesized product.
CD BioGlyco delivered our complex PNA-CPP conjugate ahead of schedule. The purity was exceptional, and the detailed MS data confirmed the integrity of both the PNA sequence and the peptide attachment. This level of quality allowed us to immediately move forward with our in vivo delivery studies, significantly accelerating our target validation timeline for our new anti-miRNA therapeutic candidate.
— Dr. Kim, Principal Investigator, Molecular Oncology Department
We had previously struggled with low yields and purity from other vendors for our highly modified PNA-FISH probes. The team at CD BioGlyco was able to implement their advanced coupling methodology, resulting in a product that passed our internal QA standards on the first attempt. Their technical consultation on the optimal dye placement was invaluable to the success of our diagnostic panel development.
— Manager, Assay Development, Clinical Genomics Laboratory
The PNA clamping reagents we received from CD BioGlyco were instrumental in developing a highly sensitive qPCR assay for detecting a low-frequency genetic mutation. The stability and specificity of their PNA allowed us to reliably suppress the wild-type amplification, which is a critical feature of our assay's regulatory submission strategy. We were very impressed with the continuous feedback during the synthesis process.
— Senior Research Scientist, Inherited Disease Center
To overcome the challenge of cellular uptake, we specialize in advanced delivery modifications. We strongly recommend leveraging our expertise in GalNAc conjugation, a highly effective technology for receptor-mediated targeting to the liver.
For liver-targeted delivery of nucleic acid aptamers.
For liver-specific delivery of miRNA inhibitors.
For developing next-generation hepatic gene therapies.
For custom, liver-targeted delivery of PNA oligomers.
CD BioGlyco is your trusted partner for high-quality, customized PNA synthesis, offering unparalleled expertise in advanced coupling chemistry, stringent quality control, and sophisticated conjugation strategies, including the crucial GalNAc-PNA delivery service. Our commitment is to deliver research-ready and clinically viable PNA molecules that drive breakthroughs in molecular diagnostics and targeted therapeutics. Contact us!
Reference
