Increased stability and specificity through combined hybridization of peptide nucleic acid (PNA) and locked nucleic acid (LNA) to supercoiled plasmids for PNA-anchored “Bioplex” formation

Lundin, Karin E.; Hašan, Miloš; Moreno, Pedro M. D.; Törnquist, Elisabeth; Oprea, Iulian; Svahn, Mathias G.; Simonson, Ernst; Smith, C. I. Edvard

doi:10.1016/j.bioeng.2005.07.003

Cited by 18 publications

(23 citation statements)

References 29 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…From these experiments we could observe the following: (1) TACH gave a two-to fourfold higher hybridization yield of bis-PNA-plasmid complexes, for the fully matched binding sites, as compared with the previous reported method [19,28]. Moreover, when using TACH the hybridization yield is even higher as compared to any constant temperature tested (Fig.…”

Section: Increasing Specificity By Identifying a Suitable Dissociatiomentioning

confidence: 59%

“…For accurate temperature control, a PCR machine was used. The bis-PNA-plasmid hybridization condition used as a control in all the experiments was the previously most used hybridization condition: 37°C, overnight with 20 mM phosphate buffer pH 6.8, 1 mM EDTA [19,28].…”

Section: Hybridization Conditionsmentioning

confidence: 99%

“…We calculated the hybridization efficacy based on the ratio between the band corresponding to hybridized (shifted) fragment and the sum of unhybridized (unshifted) and hybridized fragment. When the hybridized fragment resulted in unclear (smear) band we calculated the hybridization efficacy as described previously [28].…”

Section: Gel Retardation Assaysmentioning

confidence: 99%

“…Binding of PNA to supercoiled plasmids has been reported before [13,15,[25][26][27] but there are only a few studies focused on the optimization of this process [19,28]. In this study we have investigated the possibility to extend the affinity and specificity window by dynamic modifications of the environmental parameters (namely, temperature and pH) during the bis-PNA-plasmid hybridization process.…”

Section: Introductionmentioning

confidence: 96%

See 3 more Smart Citations

Temperature-Assisted Cyclic Hybridization (TACH): An Improved Method for Supercoiled DNA Hybridization

et al. 2010

Self Cite

View full text Add to dashboard Cite

Accurate hybridization is dependent on the ratio between sequence-specific and unspecific binding. Dissociation of unspecifically bound, while maintaining specifically hybridized, nucleic acids are key steps to obtain a well-defined complex. We have developed a new method, temperature-assisted, cyclic hybridization (TACH), which increases cognate binding at the expense of unspecific hybridization. The method was used for optimizing binding of peptide nucleic acid (PNA) to supercoiled plasmids and has several advantages over previous methods: (1) it reduces the required amount of bis-PNA by three- to fourfold; (2) it results in less unspecific binding; (3) it extends cooperative hybridization, from 3 bp to 5 bp between two adjacent binding sites; and (4) it decreases the aggregation of bis-PNA. This method might be extended to other forms of hybridizations including the use of additional nucleic acids analogs, such as locked nucleic acid (LNA) and, also, to other areas where PNAs are used such as fluorescence in situ hybridization (FISH), microarrays, or in vivo plasmid delivery.

show abstract

Section: Increasing Specificity By Identifying a Suitable Dissociatiomentioning

confidence: 59%

Section: Hybridization Conditionsmentioning

confidence: 99%

Section: Gel Retardation Assaysmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 96%

See 2 more Smart Citations

Temperature-Assisted Cyclic Hybridization (TACH): An Improved Method for Supercoiled DNA Hybridization

et al. 2010

Self Cite

View full text Add to dashboard Cite

show abstract

“…One way to facilitate the specific interaction is to let the anchors bind to sites in close proximity to each other. Both ''linear'' PNA (Lundin et al 2004) and bisPNA (Kurakin et al 1998;Lundin et al 2005) show cooperative binding and enhanced labelling of the plasmid, if the target sites are placed within a distance of three to four bases or less. The use of partly overlapping ''opener ''molecules will further facilitate the binding of positively charged PNA-NLS fusions to the correct sites and allow the use of lower concentrations .…”

Section: Increasing Intracellular Transport and Nuclear Transfermentioning

confidence: 98%

Nanotechnology approaches for gene transfer

et al. 2009

Self Cite

View full text Add to dashboard Cite

In both basic research as well as experimental gene therapy the need to transfer genetic material into a cell is of vital importance. The cellular compartment, which is the target for the genetic material, depends upon application. An siRNA that mediates silencing is preferably delivered to the cytosol while a transgene would need to end up in the nucleus for successful transcription to occur. Furthermore the ability to regulate gene expression has grown substantially since the discovery of RNA interference. In such diverse fields as medical research and agricultural pest control, the capability to alter the genetic output has been a useful tool for pushing the scientific frontiers. This review is focused on nanotechnological approaches to assemble optimised structures of nucleic acid derivatives to facilitate gene delivery as well as promoting down regulation of endogenous genes.

show abstract

A highly sensitive PNA‐microarray system for miRNA122 recognition

Forte

Ventimiglia²,

Pesaturo

et al. 2022

Biotechnology Journal

View full text Add to dashboard Cite

Surface chemistry is a fundamental aspect of the development of the sensitive biosensor based on microarray technology. Here, an advanced PNA‐microarray system for the detection of miRNA, composed by a multilayered Si/Al/Agarose component, is described. A straightforward optical signal enhancement is achieved thanks to a combination of the Al film mirror effect and the positive interference for the emission wavelength of the Cy5 fluorescent label tuned by the agarose film. The PNA‐microarray was investigated for the detection of miRNA_122, resulting in a sensitivity of about 1.75 μM–1 and Limit of Detection in the range of 0.043 nM as a function of the capture probe sequence. The contribution, in terms of H‐bonds amounts at 298 and 333 K, of the agarose coating to the dsPNA‐RNA interactions was demonstrated by Molecular Dynamic simulations. These results pave the way for advanced sensing strategies suitable for the environmental monitoring and the public safety.

show abstract

Increased stability and specificity through combined hybridization of peptide nucleic acid (PNA) and locked nucleic acid (LNA) to supercoiled plasmids for PNA-anchored “Bioplex” formation

Cited by 18 publications

References 29 publications

Temperature-Assisted Cyclic Hybridization (TACH): An Improved Method for Supercoiled DNA Hybridization

Temperature-Assisted Cyclic Hybridization (TACH): An Improved Method for Supercoiled DNA Hybridization

Nanotechnology approaches for gene transfer

A highly sensitive PNA‐microarray system for miRNA122 recognition

Contact Info

Product

Resources

About