Molecular insights on the dynamic stability of peptide nucleic acid functionalized carbon and boron nitride nanotubes.

The strategic approaches to the design of self-assembled hybrids of biomolecular systems at the nanoscale such as deoxyribonucleic acid (DNA) with single-wall carbon nanotubes (CNTs) and their structural analog, boron nitride nanotubes (BNNTs), rely on understanding how biomolecules recognize and mediate intermolecular interactions with the nanomaterial's surface. In this paper, we consider peptide nucleic acid (PNA), which is a synthetic analog of DNA, and investigate its interaction with a zigzag CNT and BNNT of similar diameter. The results based on the molecular dynamics method find that PNA provides definitive contrasts in the adsorption on the tubular surface in aqueous solution: it prefers to wrap along the circumferential direction on a (11,0) CNT, whereas it binds along the axial direction adopting an extended configuration on a (11,0) BNNT. Moreover, gas-phase Monte Carlo simulations show a dependence of the nanotube diameter on the calculated adsorption energy, with BNNTs exhibiting higher adsorption energy compared to CNTs, and the largest-diameter (25,0) tubular configuration facilitates encapsulation of PNA rather than PNA being adsorbed on its sidewall. The results are expected to be of relevance in the design of novel PNA-based archetypal hybrid materials for nanoscale applications in health-related areas including biosensing.

A rapid bioanalytical tool for detection of sequence-specific circular DNA and mitochondrial DNA point mutations.

Mutations in mitochondrial DNA (mtDNA) have been an essential cause of numerous diseases, making their identification critically important. The majority of mtDNA screening techniques require polymerase chain reaction (PCR) amplification, enzymatic digestion, and denaturation procedures, which are laborious and costly. Herein, we developed a sensitive PCR-free electrokinetic-based sensor combined with a customized bis-peptide nucleic acid (bis-PNA) and gamma-PNA (γ-PNA) probes immobilized on beads, for the detection of mtDNA point mutations and sequence-specific supercoiled plasmid DNA at the picomolar range. The probes are capable of invading the double-stranded circular DNA and forming a stable triplex structure. Thus, this method can significantly reduce the sample preparation and omit the PCR amplification steps prior to sensing. Further, this bioanalytical tool can open up a new paradigm in clinical settings for the screening of double-stranded circular nucleic acids with a single-base mismatch specificity in a rapid and sensitive manner.

Amplification-free, sequence-specific 16S rRNA detection at 1 aM.

A nucleic acid amplification-free, optics-free platform has been demonstrated for sequence-specific detection of Escherichia coli (E. coli) 16S rRNA at 1 aM (10-18 M) against a 106-fold (1 pM) background of Pseudomonas putida (P. putida) RNA. This work was driven by the need for simple, rapid, and low cost means for species-specific bacterial detection at low concentration. Our simple, conductometric sensing device functioned by detecting blockage of a nanopore fabricated in a sub-micron-thick glass membrane. Upon sequence-specific binding of target 16S rRNA, otherwise charge-neutral, PNA oligonucleotide probe-polystyrene bead conjugates become electrophoretically mobile and are driven to the glass nanopore of lesser diameter, which is blocked, thereby generating a large, sustained and readily observable step decrease in ionic current. No false positive signals were observed with P. putida RNA when this device was configured to detect E. coli 16S rRNA. Also, when a universal PNA probe complementary to the 16S rRNA of both E. coli and P. putida was conjugated to beads, a positive response to rRNA of both bacterial species was observed. Finally, the device readily detected E. coli at 10 CFU mL-1 in a 1 mL sample, also against a million-fold background of viable P. putida. These results suggest that this new device may serve as the basis for small, portable, low power, and low-cost systems for rapid detection of specific bacterial species in clinical samples, food, and water.

Prognostic significance of The Wilms' Tumor-1 (WT1) rs16754 polymorphism in acute myeloid leukemia.

Acute myeloid leukemia is a genetically heterogeneous disease characterized by the accumulation of mutations in hematopoietic progenitor cells. For its heterogeneity, prognostic markers are very useful for therapeutic choice. The most important prognostic markers are age, white blood cell count, chromosomal alterations and gene mutations. Recent works have studied the prognostic significance of WT1 polymorphisms and mutations, highlighting the role of SNP rs16754 as a positive prognostic factor in AML patients. Nevertheless, the data are still unclear. To investigate the role of WT1 rs16754 polymorphism in AML, we designed a new tool for the detection using PNA directed PCR Clamping technology. Our data were able to establish a correlation between SNP rs16754 and the clinical outcome. Our results support the hypothesis that rs16754 polymorphism is an independent positive prognostic molecular marker that could be useful for therapeutic choice. In view of this, we described a novel assay faster, more sensitive and cheaper than DNA sequencing. The assay allows evaluating WT1 rs16754 polymorphism in diagnostic routine to improve prognostic information faster and without over-costing for diagnostic laboratories.

Facile access to modified and functionalized PNAs through Ugi-based solid phase oligomerization.

Peptide nucleic acids (PNAs) derivatized with functional molecules are increasingly used in diverse biosupramolecular applications. PNAs have proven to be highly tolerant to modifications and different applications benefit from the use of modified PNAs, in particular modifications at the γ position. Herein we report simple protocols to access modified PNAs from iterative Ugi couplings which allow modular modifications at the α, ß or γ position of the PNA backbone from simple starting materials. We demonstrate the utility of the method with the synthesis of several bioactive small molecules (a peptide ligand, a kinase inhibitor and a glycan)-PNA conjugates.

In situ aneuploidy assessment in human sperm: the use of primed in situ and peptide nucleic acid-fluorescence in situ hybridization techniques.

Both the primed in situ (PRINS) and the peptide nucleic acid-fluorescence in situ hybridization (PNA-FISH) techniques constitute alternatives to the conventional (fluorescence in situ hybridization, FISH) procedure for chromosomal investigations. The PRINS reaction is based on the use of a DNA polymerase and labeled nucleotide in an in situ primer extension reaction. Peptide nucleic acid probes are synthetic DNA analogs with uncharged polyamide backbones. The two procedures present several advantages (specificity, rapidity and discriminating ability) that make them very attractive for cytogenetic purposes. Their adaptation to human spermatozoa has allowed the development of new and fast procedures for the chromosomal screening of male gametes and has provided efficient complements to FISH for in situ assessment of aneuploidy in male gametes.