Zip Nucleic Acids (ZNA)

Zip nucleic acids (ZNAs) are oligonucleotides conjugated with cationic spermine units (1). These units increase affinity of the ZNA oligo to its target by decreasing electrostatic repulsion between negatively charged anionic single strand nucleic acids. This results in improved hybridization, thus enhancing and accelerating target recognition (1,2).

The possibility of modulating the global charge of the ZNA oligonucleotide by the number of cationic spermine moieties attached to the nucleic acid oligomer is key to easily predict the melting temperature of ZNA-DNA/ZNA-RNA hybrids. Tm increases linearly with the length of the oligocation (1).

ZNAs enable specific and sensitive reactions when used as primers for PCR and Reverse Transcription (3). Moreover, ZNA probes provide broad flexibility in assay design and represent an effective alternative to Peptide-nucleic acids (PNA), Minor Groove Binder (MGB)-, Locked Nucleic Acid (LNA)-containing oligonucleotides (3,4).

The relative Tm increasing effect of ZNA or other duplex stabilizing groups is naturally higher on shorter oligonucleotides used as primers or probes. Therefore, metabion offers ZNA length ranges starting from 8mers for primers and 10mers for dual labeled probes!

The approximate Tm of ZNA® can be calculated applying the following equation:

Tm (ZNA®) = Tm (DNA) + 36z/(N-3.2)

z: number of cationic units

N: number of nucleotides

Example:

Sequence ATATATAT 8mer Tm(DNA) = 16°C

plus

ZNA-2 building block

Tm(ZNA®) = 16 + 36*2/(8-3.2) = 31°C

ZNA-2 modification almost doubles Tm; Tm increase of approx 15°C!

The number of cationic spermines is a key factor in the oligo design, as a high number of cationic spermines may result in a global charge shift of the oligonucleotide and consequent solubility issues. Metabion offers ZNA-2 and ZNA-3 (cationic) building blocks for (anionic) primers ranging from 8 to 15mers, and dual labeled probes ranging from 10 to 17mers, respectively. Attachment of ZNA-4 and ZNA-5 building blocks to primers is allowed from 16mers (ZNA-4), and 20mers (ZNA-5), respectively. Attachment of ZNA-4 and ZNA-5 building blocks to dual labeled probes is allowed from 18mers (ZNA-4), and 22mers (ZNA-5), respectively.

Additional affinity-to-target enhancement of primers and probes while maintaining specificity can be provided by incorporation of our base analogues

C-5 propynyl-dC (pdC) raising melting temperature by ~2.8°C per substitution

and/or

C-5 propynyl-dU (pdU) raising melting temperature by ~1.7°C per substitution.

On top of this, metabion also offers 5´ZNA modified dual labeled probes, introducing certain 5´reporters (i. e. FAM, Fluo, JOE, ROX, TAMRA, CY3, CY5, CY5.5, …) through “click” chemistry (click here to learn more about click-chemistry). These probes are not degraded during PCR. Therefore, is appropriately designed, they allow for high resolution, post-PCR melt curve analysis. Please inquire at info@metabion.com.

ZNA oligonucleotides have been applied successfully for a variety of applications, including antisense technology and in-situ hybridization. Visit our Special Approaches and Solution Portfolio for information about how we can assist you with your experiments!

ZNA1

ZNA2

ZNA3

ZNA4

References and links

1) Noir, R. et al. (2008) Oligonucleotide-oligospermine conjugates (Zip Nucleic Acids): a convenient means of finely tuning hybridization temperatures. J Am Chem Soc, 130, 13500-13505. Download Article

2) Voirin, E. et al. (2007) Versatile synthesis of oligodeoxyribonucleotide-oligospermine conjugates. Nat Protoc, 2, 1360-1367. Download Article

3) Moreau et al. (2009) Zip nucleic acids (ZNAs): new high affinity oligonucleotides as potent primers for PCR and reverse transcription. Nucl. Acids Res., 37: e130; doi:10.1093/nar/gkp661. Download Article

4) Paris, C. et al. (2010) Zip nucleic acids are potent hydrolysis probes for quantitative PCR.Nucl. Acids Res., doi: 10.1093/nar/gkp1218. Download Article

5) ZNA®-FAQs

6) ZNA products are sold for research use only. Please refer to our “Trademarks and License” webpage for more information on Polyplus-transfection licensing policy.