Heiko Kuhn, Bichismita Sahu, Srinivas Rapireddy, Danith H Ly, Maxim D Frank-Kamenetskii
{"title":"Sequence specificity at targeting double-stranded DNA with a γ-PNA oligomer modified with guanidinium G-clamp nucleobases.","authors":"Heiko Kuhn, Bichismita Sahu, Srinivas Rapireddy, Danith H Ly, Maxim D Frank-Kamenetskii","doi":"10.4161/adna.1.1.12444","DOIUrl":"https://doi.org/10.4161/adna.1.1.12444","url":null,"abstract":"<p><p>γ-PNA, a new class of peptide nucleic acids, promises to overcome previous sequence limitations of double-stranded DNA (dsDNA) targeting with PNA. To check the potential of γ-PNA, we have synthesized a biotinylated, pentadecameric γ-PNA of mixed sequence carrying three guanidinium G-clamp nucleobases. We have found that strand invasion reactions of the γ-PNA oligomer to its fully complementary target within dsDNA occurs with significantly higher binding rates than to targets containing single mismatches. Association of the PNA oligomer to mismatched targets does not go to completion but instead reaches a stationary level at or below 60%, even at conditions of very low ionic strength. Initial binding rates to both matched and mismatched targets experience a steep decrease with increasing salt concentration. We demonstrate that a linear DNA target fragment with the correct target sequence can be purified from DNA mixtures containing mismatched target or unrelated genomic DNA by affinity capture with streptavidin-coated magnetic beads. Similarly, supercoiled plasmid DNA is obtained with high purity from an initial sample mixture that included a linear DNA fragment with the fully complementary sequence. Based on the results obtained in this study we believe that γ-PNA has a great potential for specific targeting of chosen duplex DNA sites in a sequence-unrestricted fashion.</p>","PeriodicalId":8444,"journal":{"name":"Artificial DNA: PNA & XNA","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2010-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.4161/adna.1.1.12444","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29947289","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Direct observation of two cyclohexenyl (CeNA) ring conformations in duplex DNA.","authors":"Koen Robeyns, Piet Herdewijn, Luc Van Meervelt","doi":"10.4161/adna.1.1.10952","DOIUrl":"https://doi.org/10.4161/adna.1.1.10952","url":null,"abstract":"<p><p>Cyclohexene Nucleic Acids (CeNA), in which the 2'-deoxyribofuranose ring of the DNA building blocks is substituted by a cyclohexenyl ring, were designed as potential mimics of natural nucleic acids for antisense and, later, for siRNA applications. CeNA units, in contrast to HNA (hexitol nucleic acid) building blocks, show more flexibility at the level of the C2'-C3' bond due to the possibility of the cyclohexenyl moiety to adopt different conformations. In order to analyze the influence of CeNA residues onto the helix conformation and hydration of natural nucleic acid structures and to verify the cyclohexenyl ring conformation, a cyclohexenyl-thymine building block was incorporated into the non-self-complementary sequence d(GCG(xT)GCG)/d(CGCACGC) with (xT) a cyclohexene residue. The crystal structure of this sequence has been determined to a resolution of 1.17 Å and contains two duplexes in the asymmetric unit. The global helices belong to the B-type family and the conformations of the cyclohexenyl rings in both duplexes are different. The cyclohexene ring adopts as well the (2)H(3)-conformation (similar to C2'-endo) as the (3)H(2)-conformation (similar to C3'-endo). The crystal packing is stabilized by cobalt hexamine residues and triplet formation.</p>","PeriodicalId":8444,"journal":{"name":"Artificial DNA: PNA & XNA","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2010-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.4161/adna.1.1.10952","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29947995","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Natural - synthetic - artificial!","authors":"Peter E Nielsen","doi":"10.4161/adna.1.1.12934","DOIUrl":"https://doi.org/10.4161/adna.1.1.12934","url":null,"abstract":"<p><p>The terms \"natural,\" \"synthetic\" and \"artificial\" are discussed in relation to synthetic and artificial chromosomes and genomes, synthetic and artificial cells and artificial life.</p>","PeriodicalId":8444,"journal":{"name":"Artificial DNA: PNA & XNA","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2010-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.4161/adna.1.1.12934","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29947291","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Irina Smolina, Nancy S Miller, Maxim Frank-Kamenetskii
{"title":"PNA-based microbial pathogen identification and resistance marker detection: an accurate, isothermal rapid assay based on genome-specific features.","authors":"Irina Smolina, Nancy S Miller, Maxim Frank-Kamenetskii","doi":"","DOIUrl":"","url":null,"abstract":"<p><p>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.</p>","PeriodicalId":8444,"journal":{"name":"Artificial DNA: PNA & XNA","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2010-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2953854/pdf/nihms-232338.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29361450","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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