A strategy for the enrichment of a DNA template that encodes a functionalized PNA oligomer is discussed. The method relies on iterated cycles of chemical translation (of the template into PNA), selection (for function), and amplification (of the survivors). Potential restrictions and future perspectives are considered.
Small RNAs hit a new target: modulation of gene expression by targeting the non-coding sequences downstream from a gene
The University of Texas researchers have recently discovered that small synthetic RNAs (sRNAs) that are complementary to sequences located 3'-outside of genes can efficiently modulate gene expression. These new findings significantly expand the transcription-regulatory potential of sRNAs, and they also may provide useful leads for other artificial nucleobase oligomers to target sequences beyond the 3′ termini of mRNA.
Adding mRNA to the list of spatially organized components in bacteria
Using LNA in situ hybridization, selected mRNAs have been shown to be spatially constrained to their chromosomal loci in two distantly related bacterial organisms. Translating ribosomes are diffusion limited by mRNA association.
1,4-linked 1,2,3-Triazole containing chimeric PNA (aeg-TzNA) oligomers: In-situ synthesis by solid phase "click" reaction and binding studies with DNA
This is the first report of synthesis of triazole analogues of PNA in which the amide bond of PNA is replaced by 1,2,3-triazole ring. The triazole ring is synthesized by click chemistry on solid resin, which is also new and novel. Triazole polyners are increasingly attracting attention recently (see introduction) and the peptide bond replacement with triazole is also of recent interest. triazole ring brings more order into the backbone upto some degree of replacement and beyond which, it becomes so self ordered that the completely modified triazole oligomers do not hybridize to complementary DNA. However the high degree of self ordering is indicated by higher thermal melting of single stranded oligomers. This is perhaps due to the higher dipole moment of triazole ring compared to amide bonds. Although we have provided data on only triplex melting of oligo triazolyl thymines, work is under progress for similar studies with duplexes. The work has lead to new class of chimeric PNA - TzNA ologomers with novel self ordered properties
PNA-based microbial pathogen identification and resistance marker detection: an accurate, isothermal rapid assay based on genome-specific features
With the rapidly growing availability of the entire genome sequences of microbial pathogens, there is unmet need for increasingly sensitive systems to monitor the gene-specific markers for diagnosis of bacteremia that enables an earlier detection of causative agent and determination of drug resistance. To address these challenges, a novel FISH-type genomic sequence-based molecular technique is proposed that can identify bacteria and simultaneously detect antibiotic resistance markers for rapid and accurate testing of pathogens. The approach is based on a synergistic combination of advanced Peptide Nucleic Acid (PNA)-based technology and signal-enhancing Rolling Circle Amplification (RCA) reaction to achieve a highly specific and sensitive assay. A specific PNA-DNA construct serves as an exceedingly selective and very effective biomarker, while RCA enhances detection sensitivity and provide with a highly multiplexed assay system. Distinct-color fluorescent decorator probes are used to identify about 20-nucleotide-long signature sequences in bacterial genomic DNA and/or key genetic markers of drug resistance in order to identify and characterize various pathogens. The technique's potential and its utility for clinical diagnostics are illustrated by identification of S. aureus with simultaneous discrimination of methicillin-sensitive (MSSA) versus methicillin-resistant (MRSA) strains. Overall these promising results hint to the adoption of PNA-based rapid sensitive detection for diagnosis of other clinically relevant organisms. Thereby, new assay enables significantly earlier administration of appropriate antimicrobial therapy and may, thus have a positive impact on the outcome of the patient.
A pyrenyl-PNA probe for DNA and RNA recognition: fluorescence and UV absorption studies
The design and the synthesis of a PNA oligomer containing a pyrenyl residue in the backbone were performed. PNA sequence was chosen complementary to a "G rich" target sequence involved in G- quadruplex formation. The pyrenyl unit replaced a nucleobase in the middle of the PNA through covalent linkage to the backbone by a carboxymethyl unit. A systematic study on the binding properties of this probe towards DNA and RNA complementary strands was carried out by UV and fluorescence spectroscopy. UV melting curves indicated that the PNA probe bind more tightly to RNA rather than to DNA. Thermodynamic data obtained by Van't Hoff fitting of the melting curves indicated that, in the case of RNA, a more favorable interaction occurs between the pyrenyl unit and the RNA nucleobases, leading to a very favorable enthalpic contribution.
The fluorescence analysis showed specific quenching of the pyrene emission associated to the formation of the full-match PNA-DNA or PNA-RNA duplexes. Again, this behavior was more evident in the case of RNA, consistent with the stronger interaction of the pyrenyl unit with the complementary strand. In order to study the sequence specificity of the pyrenyl-PNA probe (pyr-PNA), recognition experiments on mismatched DNA and RNA sequences were also performed.
Hydrogelation abilities of nucleobase-modified cytidines possessing substituted triazoles
Nucleoside-derived hydrogelators have been sought for their potential biomedical applications, such as are found in tissue engineering and drug delivery. By judiciously adding a degree of hydrophobicity certain analogues are able to form micelles, bi-layers and gels in water. Research in this area has yet to lay down solid ground rules for the rational design of novel nucleoside gelators making further studies necessary. The synthesis and examination of a series of aryl-substituted 5-triazolylcytidines yielded an analogue that gelates water. 5-(1-(2,2'-bithiophen-3-yl)-1H-1,2,3-triazol-4-yl)-2'-deoxycytidine was found to form gels in water down to 0.3 wt%. The ribocytidine analogue failed to form gel in aqueous solution; but was able to form a hydrogel in the presence of guanosine. Images obtained by SEM show the different architectures of the gel; varying from cribriform, to fibrous, to lamellar. The present gelating compound studied may have potential as a component of a controlled-release drug delivery system.