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Utilising the left-helical conformation of L-DNA for analysing different marker types on a single universal microarray platform.

Hauser NC, Martinez R, Jacob A, Rupp S, Hoheisel JD, Matysiak S - Nucleic Acids Res. (2006)

Bottom Line: Because of its chiral difference, L-DNA does not bind to its naturally occurring D-DNA counterpart, however.Typical results for the measurement of transcript level variations, genotypic differences and DNA-protein interactions are presented.However, on the basis of the characteristic features of L-DNA, also other applications of this molecule type are discussed.

View Article: PubMed Central - PubMed

Affiliation: Genomics-Proteomics-Systemsbiology, Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik Nobelstrasse 12, 70569 Stuttgart, Germany. nicole.hauser@igb.fraunhofer.de

ABSTRACT
L-DNA is the perfect mirror-image form of the naturally occurring d-conformation of DNA. Therefore, L-DNA duplexes have the same physical characteristics in terms of solubility, duplex stability and selectivity as D-DNA but form a left-helical double-helix. Because of its chiral difference, L-DNA does not bind to its naturally occurring D-DNA counterpart, however. We analysed some of the properties that are typical for L-DNA. For all the differences, L-DNA is chemically compatible with the D-form of DNA, so that chimeric molecules can be synthesized. We take advantage of the characteristics of L-DNA toward the establishment of a universal microarray that permits the analysis of different kinds of molecular diagnostic information in a single experiment on a single platform, in various combinations. Typical results for the measurement of transcript level variations, genotypic differences and DNA-protein interactions are presented. However, on the basis of the characteristic features of L-DNA, also other applications of this molecule type are discussed.

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Hybridization specificity of L-DNA and D-DNA. Identical sets of 102 probe oligonucleotides were synthesized in both D- and L-form and spotted next to each other onto microarrays. They were subjected to hybridizations with a set of 30 fluorescently labelled complementary oligonucleotides made of either L-DNA (a and c) or D-DNA (b and d). The raw signal intensities (in arbitrary units) obtained at the 204 spot positions are shown here. Each set of hybridized oligomers is complementary to the probes represented by the orange area. While cross-hybridization occurred within each enantiomeric class at low-stringency (45°C; a and b), there was none between different enantiomers. As expected, cross-hybridization was much reduced at high-stringency conditions (65°C; c and d). Variation in signal intensities also accounts for differences in the amounts of oligomer attached to the support at the various spot positions.
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fig3: Hybridization specificity of L-DNA and D-DNA. Identical sets of 102 probe oligonucleotides were synthesized in both D- and L-form and spotted next to each other onto microarrays. They were subjected to hybridizations with a set of 30 fluorescently labelled complementary oligonucleotides made of either L-DNA (a and c) or D-DNA (b and d). The raw signal intensities (in arbitrary units) obtained at the 204 spot positions are shown here. Each set of hybridized oligomers is complementary to the probes represented by the orange area. While cross-hybridization occurred within each enantiomeric class at low-stringency (45°C; a and b), there was none between different enantiomers. As expected, cross-hybridization was much reduced at high-stringency conditions (65°C; c and d). Variation in signal intensities also accounts for differences in the amounts of oligomer attached to the support at the various spot positions.

Mentions: In order to analyse the specificity of the interaction between L-DNA and D-DNA, an identical set of 102 ZIP-code oligonucleotides of 20 nt in length was synthesized in either conformation (see Supplementary Table S1). In addition, we produced 30 fully complementary oligonucleotides in both l- and d-conformation that were labelled at their 5′-terminus with a fluorescence dye. The D- and L-form ZIP-code libraries were spotted next to each other onto microarray surfaces and subjected to hybridizations with the respective set of complementary fluorescently labelled sequences (Figure 3). Even at conditions of low stringency that produced an enormous amount of cross-hybridization of the 30 labelled oligonucleotides at the ZIP-code sequences of the same conformation, no signal above background could be observed at the spots at which the ZIP-code library of the opposite conformation had been placed. As expected, the degree of cross-hybridization within each enantiomeric class decreased with increasing stringency of the hybridization conditions.


Utilising the left-helical conformation of L-DNA for analysing different marker types on a single universal microarray platform.

Hauser NC, Martinez R, Jacob A, Rupp S, Hoheisel JD, Matysiak S - Nucleic Acids Res. (2006)

Hybridization specificity of L-DNA and D-DNA. Identical sets of 102 probe oligonucleotides were synthesized in both D- and L-form and spotted next to each other onto microarrays. They were subjected to hybridizations with a set of 30 fluorescently labelled complementary oligonucleotides made of either L-DNA (a and c) or D-DNA (b and d). The raw signal intensities (in arbitrary units) obtained at the 204 spot positions are shown here. Each set of hybridized oligomers is complementary to the probes represented by the orange area. While cross-hybridization occurred within each enantiomeric class at low-stringency (45°C; a and b), there was none between different enantiomers. As expected, cross-hybridization was much reduced at high-stringency conditions (65°C; c and d). Variation in signal intensities also accounts for differences in the amounts of oligomer attached to the support at the various spot positions.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC1636439&req=5

fig3: Hybridization specificity of L-DNA and D-DNA. Identical sets of 102 probe oligonucleotides were synthesized in both D- and L-form and spotted next to each other onto microarrays. They were subjected to hybridizations with a set of 30 fluorescently labelled complementary oligonucleotides made of either L-DNA (a and c) or D-DNA (b and d). The raw signal intensities (in arbitrary units) obtained at the 204 spot positions are shown here. Each set of hybridized oligomers is complementary to the probes represented by the orange area. While cross-hybridization occurred within each enantiomeric class at low-stringency (45°C; a and b), there was none between different enantiomers. As expected, cross-hybridization was much reduced at high-stringency conditions (65°C; c and d). Variation in signal intensities also accounts for differences in the amounts of oligomer attached to the support at the various spot positions.
Mentions: In order to analyse the specificity of the interaction between L-DNA and D-DNA, an identical set of 102 ZIP-code oligonucleotides of 20 nt in length was synthesized in either conformation (see Supplementary Table S1). In addition, we produced 30 fully complementary oligonucleotides in both l- and d-conformation that were labelled at their 5′-terminus with a fluorescence dye. The D- and L-form ZIP-code libraries were spotted next to each other onto microarray surfaces and subjected to hybridizations with the respective set of complementary fluorescently labelled sequences (Figure 3). Even at conditions of low stringency that produced an enormous amount of cross-hybridization of the 30 labelled oligonucleotides at the ZIP-code sequences of the same conformation, no signal above background could be observed at the spots at which the ZIP-code library of the opposite conformation had been placed. As expected, the degree of cross-hybridization within each enantiomeric class decreased with increasing stringency of the hybridization conditions.

Bottom Line: Because of its chiral difference, L-DNA does not bind to its naturally occurring D-DNA counterpart, however.Typical results for the measurement of transcript level variations, genotypic differences and DNA-protein interactions are presented.However, on the basis of the characteristic features of L-DNA, also other applications of this molecule type are discussed.

View Article: PubMed Central - PubMed

Affiliation: Genomics-Proteomics-Systemsbiology, Fraunhofer-Institut für Grenzflächen- und Bioverfahrenstechnik Nobelstrasse 12, 70569 Stuttgart, Germany. nicole.hauser@igb.fraunhofer.de

ABSTRACT
L-DNA is the perfect mirror-image form of the naturally occurring d-conformation of DNA. Therefore, L-DNA duplexes have the same physical characteristics in terms of solubility, duplex stability and selectivity as D-DNA but form a left-helical double-helix. Because of its chiral difference, L-DNA does not bind to its naturally occurring D-DNA counterpart, however. We analysed some of the properties that are typical for L-DNA. For all the differences, L-DNA is chemically compatible with the D-form of DNA, so that chimeric molecules can be synthesized. We take advantage of the characteristics of L-DNA toward the establishment of a universal microarray that permits the analysis of different kinds of molecular diagnostic information in a single experiment on a single platform, in various combinations. Typical results for the measurement of transcript level variations, genotypic differences and DNA-protein interactions are presented. However, on the basis of the characteristic features of L-DNA, also other applications of this molecule type are discussed.

Show MeSH
Related in: MedlinePlus