共查询到20条相似文献,搜索用时 15 毫秒
1.
Jesper Wengel Michael Petersen Miriam Frieden Troels Koch 《International journal of peptide research and therapeutics》2005,10(3):237-253
Summary Preparation of LNA nucleosides requires a number of synthetic steps but very efficient procedures have been developed, as have protocols for synthesis of LNA oligonucleotides on automated DNA synthesizers. In all cases, LNA oligonucleotides have exhibited good aqueous solubility as would be expected from their close structural resemblance to the natural nucleic acids. The universality of LNA mediated high-affinity and specific hybridization has been demonstrated extensively with a large number of duplex forming LNA-oligonucleotides. Most importantly, most of the members of the LNA molecular family have been shown to exert their substantial affinity increase (i) in combination with standard DNA, RNA and contemporary analogues and (ii) whether inserted as single nucleosides in an oligonucleotide or as blocks of contiguous nucleotides, an important point. The works on TFOs is expanding the usefulness of LNA to double strand recognition and it has been demonstrated that LNA it is a promising structure for further base modifications in the pursuit of global sequence specific recognition of DNA. 相似文献
2.
Jesper Wengel Michael Petersen Miriam Frieden Troels Koch 《Letters in Peptide Science》2003,10(3-4):237-253
Preparation of LNA nucleosides requires a number of synthetic steps but very efficient procedures have been developed, as
have protocols for synthesis of LNA oligonucleotides on automated DNA synthesizers. In all cases, LNA oligonucleotides have
exhibited good aqueous solubility as would be expected from their close structural resemblance to the natural nucleic acids.
The universality of LNA mediated high-affinity and specific hybridization has been demonstrated extensively with a large number
of duplex forming LNA-oligonucleotides. Most importantly, most of the members of the LNA molecular family have been shown
to exert their substantial affinity increase (i) in combination with standard DNA, RNA and contemporary analogues and (ii)
whether inserted as single nucleosides in an oligonucleotide or as blocks of contiguous nucleotides, an important point. The
works on TFO's is expanding the usefulness of LNA to double strand recognition and it has been demonstrated that LNA it is
a promising structure for further base modifications in the pursuit of global sequence specific recognition of DNA. 相似文献
3.
Jesper Wengel Michael Petersen Miriam Frieden Troels Koch 《International journal of peptide research and therapeutics》2003,10(3-4):237-253
Summary Preparation of LNA nucleosides requires a number of synthetic steps but very efficient procedures have been developed, as
have protocols for synthesis of LNA oligonucleotides on automated DNA synthesizers. In all cases, LNA oligonucleotides have
exhibited good aqueous solubility as would be expected from their close structural resemblance to the natural nucleic acids.
The universality of LNA mediated high-affinity and specific hybridization has been demonstrated extensively with a large number
of duplex forming LNA-oligonucleotides. Most importantly, most of the members of the LNA molecular family have been shown
to exert their substantial affinity increase (i) in combination with standard DNA, RNA and contemporary analogues and (ii)
whether inserted as single nucleosides in an oligonucleotide or as blocks of contiguous nucleotides, an important point. The
works on TFO's is expanding the usefulness of LNA to double strand recognition and it has been demonstrated that LNA it is
a promising structure for further base modifications in the pursuit of global sequence specific recognition of DNA. 相似文献
4.
A 3'-C-branched LNA-type bicyclic nucleoside, containing a furanose ring locked in an N-type conformation, was synthesized from a known 3-C-vinyl allofuranose derivative using a strategy relying on the condensation with the nucleobase after the introduction of the branching hydroxymethyl chain by our recently developed RuO4 based protocol. This branched LNA nucleoside has a potential as a monomer for the functionalization of LNA. 相似文献
5.
A sensitive array for microRNA expression profiling (miChip) based on locked nucleic acids (LNA) 总被引:23,自引:3,他引:20
Castoldi M Schmidt S Benes V Noerholm M Kulozik AE Hentze MW Muckenthaler MU 《RNA (New York, N.Y.)》2006,12(5):913-920
MicroRNAs represent a class of short (approximately 22 nt), noncoding regulatory RNAs involved in development, differentiation, and metabolism. We describe a novel microarray platform for genome-wide profiling of mature miRNAs (miChip) using locked nucleic acid (LNA)-modified capture probes. The biophysical properties of LNA were exploited to design probe sets for uniform, high-affinity hybridizations yielding highly accurate signals able to discriminate between single nucleotide differences and, hence, between closely related miRNA family members. The superior detection sensitivity eliminates the need for RNA size selection and/or amplification. MiChip will greatly simplify miRNA expression profiling of biological and clinical samples. 相似文献
6.
Nielsen CB Singh SK Wengel J Jacobsen JP 《Journal of biomolecular structure & dynamics》1999,17(2):175-191
LNA (Locked Nucleic Acids) is a novel oligonucleotide analogue containing a conformationally restricted nucleotide with a 2'-O, 4'-C-methylene bridge that induces unprecedented thermal affinities when mixed with complementary single stranded DNA and RNA. We have used two-dimensional 1H NMR spectroscopy obtained at 750 and 500 MHz to determine a high resolution solution structure of an LNA oligonucleotide hybridized to the complementary DNA strand. The determination of the structure was based on a complete relaxation matrix analysis of the NOESY cross peaks followed by restrained molecular dynamics calculations. Forty final structures were generated for the duplex from A-type and B-type dsDNA starting structures. The root-mean-square deviation (RMSD) of the coordinates for the forty structures of the complex was 0.32A. The structures were analysed by use of calculated helix parameters. This showed that the values for rise and buckle in the LNA duplex is markedly different from canonical B-DNA at the modification site. A value of twist similar to A-DNA is also observed at the modification site. The overall length of the helix which is 27.3 A. The average twist over the sequence are 35.9 degrees +/- 0.3 degrees. Consequently, the modification does not cause the helix to unwind. The bis-intercalation of the thiazole orange dye TOTO to the LNA duplex was also investigated by 1H NMR spectroscopy to sense the structural change from the unmodified oligonucleotide. We observed that the bis-intercalation of TOTO is much less favourable in the 5'-CT(L)AG-3' site than in the unmodified 5'-CTAG-3' site. This was related to the change in the base stacking of the LNA duplex compared to the unmodified duplex. 相似文献
7.
We have determined the NMR solution structures of the quadruplexes formed by d(TGLGLT) and d(TL4T), where L denotes LNA (locked nucleic acid) modified G-residues. Both structures are tetrameric, parallel and right-handed and the native global fold of the corresponding DNA quadruplex is retained upon introduction of the LNA nucleotides. However, local structural alterations are observed owing to the locked LNA sugars. In particular, a distinct change in the sugar–phosphate backbone is observed at the G2pL3 and L2pL3 base steps and sequence dependent changes in the twist between tetrads are also seen. Both the LNA modified quadruplexes have raised thermostability as compared to the DNA quadruplex. The quadruplex-forming capability of d(TGLGLT) is of particular interest as it expands the design flexibility for stable parallel LNA quadruplexes and shows that LNA nucleotides can be mixed with DNA or other modified nucleic acids. As such, LNA-based quadruplexes can be decorated by a variety of chemical modifications. Such LNA quadruplex scaffolds might find applications in the developing field of nanobiotechnology. 相似文献
8.
Locked Nucleic Acid (LNA) is a novel, third generation DNA analogue that has the potentialto impact strongly on the future
development of a diversity of nucleic acid based technologies.The present chapter reviews the known biochemical properties
of LNA and exemplifies how thesehave been used to improve both DNA diagnostic technologies and antisense therapeutics. 相似文献
9.
Henrik Ørum Andreas Wolter Lars Kongsbak 《International journal of peptide research and therapeutics》2003,10(3-4):325-334
Summary Locked Nucleic Acid (LNA) is a novel, third generation DNA analogue that has the potential to impact strongly on the future
development of a diversity of nucleic acid based technologies. The present chapter reviews the known biochemical properties
of LNA and exemplifies how these have been used to improve both DNA diagnostic technologies and antisense therapeutics. 相似文献
10.
Sandra Prepens Karl-Anton Kreuzer Fabian Leendertz Andreas Nitsche Bernhard Ehlers 《Virology journal》2007,4(1):1-15
The human papillomavirus type 16 E5 protein (HPV16 E5) down-regulates surface expression of HLA-I molecules. The molecular mechanisms underlying this effect are so far unknown. Here we show that HPV16 E5 down-regulates HLA-I surface expression in calnexin-containing but not in calnexin-deficient cells. Immunoprecipitation experiments reveal that calnexin and HPV16E5 can be co-precipitated and that this association depends on the presence of a wild-type first hydrophobic region of E5. When an E5 mutant (M1) in which the first putative transmembrane helix had been disrupted was used for the transfections calnexin-E5 co-precipitation was strongly impaired. In addition, we show that the M1 mutant is only able to marginally down-regulate HLA-I surface expression compared to the wild-type protein. Besides, we demonstrate that E5 forms a ternary complex with calnexin and the heavy chain of HLA-I, which is mediated by the first hydrophobic region of the E5 protein. On the basis of our results we conclude that formation of this complex is responsible for retention of HLA-I molecules in the ER of the cells. 相似文献
11.
Henrik Ørum Andreas Wolter Lars Kongsbak 《International journal of peptide research and therapeutics》2005,10(3):325-334
Summary Locked Nucleic Acid (LNA) is a novel, third generation DNA analogue that has the potential to impact strongly on the future development of a diversity of nucleic acid based technologies. The present chapter reviews the known biochemical properties of LNA and exemplifies how these have been used to improve both DNA diagnostic technologies and antisense therapeutics. 相似文献
12.
Vollmer J Jepsen JS Uhlmann E Schetter C Jurk M Wader T Wüllner M Krieg AM 《Oligonucleotides》2004,14(1):23-31
Locked nucleic acid (LNA) is an RNA derivative that when introduced into oligodeoxynucleotides (ODN), mediates high efficacy and stability. CpG ODNs are potent immune stimulators and are recognized by toll-like receptor-9 (TLR9). Some phosphorothioate antisense ODNs bearing CpG dinucleotides have been shown to possess immune modulatory capacities. We investigated the effects of LNA substitutions on immune stimulation mediated by antisense ODN G3139 or CpG ODN 2006. LNA ODNs were tested for their ability to stimulate cytokine secretion from human immune cells or TLR9-dependent signaling. Phosphorothioate chimeric LNA/DNA antisense ODNs with phosphodiester-linked LNA nucleobases at both ends showed a marked decrease of immune modulation with an increasing number of 3' and 5' LNA bases. In addition, guanosine-LNA and cytosine-LNA or simply cytosine-LNA substitutions in the CpG dinucleotides of ODN 2006 led to strong decrease or near complete loss of immune modulation. TLR9-mediated signaling was similarly affected. These data indicate that increasing amounts of LNA residues in the flanks or substitutions of CpG nucleobases with LNA reduce or eliminate the immune stimulatory effects of CpG-containing phosphorothioate ODN. 相似文献
13.
The design of modified nucleic acid probes, primers, and therapeutics is improved by considering their thermodynamics. Locked nucleic acid (LNA) is one of the most useful modified backbones, with incorporation of a single LNA providing a substantial increase in duplex stability. In this work, the hybridization DeltaH(o), DeltaS(o), and melting temperature (T(M)) were measured from absorbance melting curves for 100 duplex oligonucleotides with single internal LNA nucleotides on one strand, and the results provided DeltaDeltaH(o), DeltaDeltaS(o), DeltaDelta, and DeltaT(M) relative to reference DNA oligonucleotides. LNA pyrimidines contribute more stability than purines, especially A(L), but there is substantial context dependence for each LNA base. Both the 5' and 3' neighbors must be considered in predicting the effect of an LNA incorporation, with purine neighbors providing more stability. Enthalpy-entropy compensation in DeltaDeltaH(o) and DeltaDeltaS(o) is observed across the set of sequences, suggesting that LNA can stabilize the duplex by either preorganization or improved stacking, but not both simultaneously. Singular value decomposition analysis provides predictive sequence-dependent rules for hybridization of singly LNA-substituted DNA oligonucleotides to their all-DNA complements. The results are provided as sets of DeltaDeltaH(o), DeltaDeltaS(o), and DeltaDelta parameters for all 32 of the possible nearest neighbors for LNA+DNA:DNA hybridization (5' MX(L) and 5' X(L)N, where M, N, and X = A, C, G, or T and X(L) represents LNA). The parameters are applicable within the standard thermodynamic prediction algorithms. They provide T(M) estimates accurate to within 2 degrees C for LNA-containing oligonucleotides, which is significantly better accuracy than previously available. 相似文献
14.
Locked nucleic acid (LNA) is a nucleic acid analogue containing one or more LNA nucleotide monomers with a bicyclic furanose unit locked in an RNA mimicking sugar conformation. LNA oligonucleotides display unprecedented hybridization affinity toward complementary single-stranded RNA and complementary single- or double-stranded DNA. Structural studies have shown that LNA oligonucleotides induce A-type (RNA-like) duplex conformations. The wide applicability of LNA oligonucleotides for gene silencing and their use for research and diagnostic purposes are documented in a number of recent reports, some of which are described herein. 相似文献
15.
Position-dependent effects of locked nucleic acid (LNA) on DNA sequencing and PCR primers 总被引:2,自引:0,他引:2
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Genomes are becoming heavily annotated with important features. Analysis of these features often employs oligonucleotides that hybridize at defined locations. When the defined location lies in a poor sequence context, traditional design strategies may fail. Locked Nucleic Acid (LNA) can enhance oligonucleotide affinity and specificity. Though LNA has been used in many applications, formal design rules are still being defined. To further this effort we have investigated the effect of LNA on the performance of sequencing and PCR primers in AT-rich regions, where short primers yield poor sequencing reads or PCR yields. LNA was used in three positional patterns: near the 5′ end (LNA-5′), near the 3′ end (LNA-3′) and distributed throughout (LNA-Even). Quantitative measures of sequencing read length (Phred Q30 count) and real-time PCR signal (cycle threshold, CT) were characterized using two-way ANOVA. LNA-5′ increased the average Phred Q30 score by 60% and it was never observed to decrease performance. LNA-5′ generated cycle thresholds in quantitative PCR that were comparable to high-yielding conventional primers. In contrast, LNA-3′ and LNA-Even did not improve read lengths or CT. ANOVA demonstrated the statistical significance of these results and identified significant interaction between the positional design rule and primer sequence. 相似文献
16.
17.
丙型肝炎病毒(HCV)是由约9.6kb个核苷酸组成的单股正链RNA病毒,属于黄病毒科丙型肝炎病毒属,被分为7个基因型及若干亚型。丙型肝炎是由HCV感染引起的常见血液传播型疾病,全球约有1.8亿人感染,引起慢性肝炎、肝硬化、肝癌。至今没有针对HCV的保护性疫苗,目前批准的标准治疗方式主要是聚乙二醇-干扰素(PEG-IFN)和利巴韦林(RBV)的联合疗法。目前这种疗法对不同基因型的感染患者的作用效果差异很大,对HCV基因2型及3型的患者,80%可以获得持续的病毒学应答反应(SVR),然而 相似文献
18.
In vivo tumor growth inhibition and biodistribution studies of locked nucleic acid (LNA) antisense oligonucleotides 总被引:4,自引:1,他引:3
Fluiter K ten Asbroek AL de Wissel MB Jakobs ME Wissenbach M Olsson H Olsen O Oerum H Baas F 《Nucleic acids research》2003,31(3):953-962
Locked nucleic acids (LNA) are novel high-affinity DNA analogs that can be used as genotype-specific drugs. The LNA oligonucleotides (LNA PO ODNs) are very stable in vitro and in vivo without the need for a phosphorothiolated backbone. In this study we tested the biological fate and the efficacy in tumor growth inhibition of antisense oligonucleotides directed against the gene of the large subunit of RNA polymerase II (POLR2A) that are completely synthesized as LNA containing diester backbones. These full LNA oligonucleotides strongly reduce POLR2A protein levels. Full LNA PO ODNs appeared to be very stable compounds when injected into the circulation of mice. Full LNA PO ODNs were continuously administered for 14 days to tumor-bearing nude mice. Tumor growth was inhibited sequence specifically at dosages from 1 mg/kg/day. LNA PO ODNs appeared to be non-toxic at dosages <5 mg/kg/day. Biodistribution studies showed the kidneys to have the highest uptake of LNA PO ODNs and urinary secretion as the major route of clearance. This report shows that LNA PO ODNs are potent genotype-specific drugs that can inhibit tumor growth in vivo. 相似文献
19.
Lauritsen A Dahl BM Dahl O Vester B Wengel J 《Bioorganic & medicinal chemistry letters》2003,13(2):253-256
Synthesis of an oligonucleotide containing one methylphosphonate locked nucleic acid (LNA) thymine monomer using the phosphoramidite approach is described. The binding affinity of this 9-mer methylphosphonate LNA towards complementary DNA and RNA oligonucleotides was increased compared to the reference DNA, but decreased compared to the reference LNA. In the 9-mer sequence context studied, introduction of a single methylphosphonate LNA monomer, contrary to a single LNA monomer, efficiently inhibits 3'-exonucleolytic degradation. 相似文献
20.
Tolstrup N Nielsen PS Kolberg JG Frankel AM Vissing H Kauppinen S 《Nucleic acids research》2003,31(13):3758-3762
We report the development of new software, OligoDesign, which provides optimal design of LNA (locked nucleic acid) substituted oligonucleotides for functional genomics applications. LNAs constitute a novel class of bicyclic RNA analogs having an exceptionally high affinity and specificity toward their complementary DNA and RNA target molecules. The OligoDesign software features recognition and filtering of the target sequence by genome-wide BLAST analysis in order to minimize cross-hybridization with non-target sequences. Furthermore it includes routines for prediction of melting temperature, self-annealing and secondary structure for LNA substituted oligonucleotides, as well as secondary structure prediction of the target nucleotide sequence. Individual scores for all these properties are calculated for each possible LNA oligonucleotide in the query gene and the OligoDesign program ranks the LNA capture probes according to a combined fuzzy logic score and finally returns the top scoring probes to the user in the output. We have successfully used the OligoDesign tool to design a Caenorhabditis elegans LNA oligonucleotide microarray, which allows monitoring of the expression of a set of 120 potential marker genes for a variety of stress and toxicological processes and toxicologically relevant pathways. The OligoDesign program is freely accessible at http://lnatools.com/. 相似文献