PCR Blockers

PNA Clamps for Host DNA Suppression in Metagenomic Studies

16S, 18S, and 28S rRNA sequencing are widely used for taxonomic classification in metagenomics. However, a frequent challenge is host-derived contamination, where the majority of sequencing reads originate from the host’s mitochondrial or plastid genomes. This can obscure the detection of low-abundance microbial taxa.

To address this, PNA clamps (PNA PCR blockers) are designed to selectively bind to host rRNA regions, suppressing their amplification and enriching microbial sequences in your dataset.

🔬 PNA PCR Blockers We Offer

Our catalog includes a range of PNA blockers targeting mitochondrial, chloroplast, and ITS DNA from multiple host species.

All products are usually in stock and available for same-day shipping for next-day delivery.

🧪 How to Use PNA PCR Blockers

• Add PNA blockers directly to the PCR mix at a final concentration of 0.5 µM to 6 µM. Temperature: 70°C – 80°C
• Include a PNA clamping step between denaturation and primer annealing.
• For optimization, tryrange of clamping temperature to 65°C–80°C (about 10°C below to 5°C above the predicted Tm of the PNA).
• Duration: 10 – 30 seconds
• Add PNA blockers directly to the PCR mix.

📥 Download the full mPNA and pPNA protocol here. 

🧬 Need a Custom Clamp?

If your host genome is not covered by the above options, we can design and synthesize a custom PNA clamp tailored to your target sequence.

📧 Contact us at info@pnabio.com for assistance, or visit our Custom PNA Oligos page for more information.

📚 Scientific References

• Practical innovations for high-throughput amplicon sequencing. Lundberg DS et al. (2013) Nat Methods 10:999-1002 (mPNA and pPNA)
• The Populus holobiont: dissecting the effects of plant niches and genotype on the microbiome. Cregger MA et al. (2018) Microbiome 6:31 (Populus mPNA and ITS PNA)
• Chloroplast sequence variation and the efficacy of peptide nucleic acids for blocking host amplification in plant microbiome studies. Fitzpatrick CR et al. (2018) Microbiome 6: 144 (Asteraceae pPNA)
• Peptide nucleic acid (PNA) clamps reduce amplification of host chloroplast and mitochondria rRNA gene sequences and increase detected diversity in 16S rRNA gene profiling analysis of oak-associated microbiota. Hussain U et al. (2025) Environmental Microbiome 20:14 (Quercus mPNA)

🧬 Overview

A validated PNA clamp that selectively suppresses amplification of the aberrant MOPC-21–derived Vκ transcript in SP2/0- or P3X63Ag8.653-based hybridomas, enabling clean amplification of the functional light-chain Vκ from your hybridoma cDNA.

🧾 Product details

• Target: MOPC-21 aberrant Vκ (CDR3 region)
• PNA sequence (antisense to abVκ CDR3): 5’ CGTGTAAGCTCCCTA
• Validated use: PNA-mediated PCR clamping to prevent abVκ amplification while preserving amplification of functional hybridoma Vκ. In the cited work, PNA clamping eliminated abVκ clones and enriched functional Vκ clones.

⚗️ PCR Reaction Mix

• Primers (e.g., Vκ2Back/Vκ4For): 0.25 µM each
• PNA clamp: 2.5~10 µM
• Template: cDNA from hybridoma
• Volume: 20–50 µL

⏱️ PCR Cycle

1.    94 °C, 2 min
2.   30 cycles of:
•  94 °C, 20 s (denaturation)
•  65 °C, 30 s (PNA preferential clamping)
•  50 °C, 30 s (primer annealing)
•  60 °C, 30 s (extension)
3.   60 °C, 2 min (final extension)

📚 Reference

• Selective PCR amplification of functional immunoglobulin light chain from hybridoma containing the aberrant MOPC 21-derived V kappa by PNA-mediated PCR clamping. Cochet O et al. (1999) Biotechniques 26(5):818-20.