Synthesis, Structure, and Deoxyribonucleic Acid Sequencing with a Universal Nucleoside: 1-(2'-Deoxy-.beta.-D-ribofuranosyl)-3-nitropyrrole

Bergstrom, Donald E.; Zhang, Peiming; Toma, Pascal H.; Andrews, Philip C.; Nichols, Robert C.

doi:10.1021/ja00109a003

Cited by 64 publications

(76 citation statements)

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“…Fluorescent label was attached to the 5′-end of oligonucleotides using 6-FAM (6-carboxyfluorescein) amidites or attached to a 3′-amino group (C3-CPG column; Glen Research, Sterling, VA) using NHS-FAM (N-hydroxysuccinimide ester of FAM; Applied Biosystems Division, Perkin-Elmer Corp). A universal nucleotide analog, 1-(2′-deoxy-β-D-ribofuranosyl)-3-nitropyrrole, herein designated Q, was synthesized, transformed to the phosphoramidite and oligonucleotides synthesized as described (28). All oligonucleotides used in this study were purified by polyacrylamide gel electrophoresis with recovery of DNA from gel slices using C-18 Sep-Pak Cartridges from Waters Division, Millipore (Bedford, MA).…”

Section: Synthesis Of Oligonucleotide Primersmentioning

confidence: 99%

“…Oligonucleotides containing the nucleotide analog 3-nitropyrrole deoxyribonucleotide (Q) at multiple sites in place of the natural nucleotides have been shown to function effectively as sequencing and PCR primers (28,42). 3-Nitropyrrole presumably allows preservation of helix integrity because it is sufficiently small to fit opposite any of the four natural bases and has high stacking potential due to a highly polarized π electronic configuration.…”

Section: Improving the Fidelity Of Tth Dna Ligasementioning

confidence: 99%

“…On the basis of modeling studies (QUANTA/CHARMM), Q can fit opposite both T and A with minimal perturbation of the helix structure (28). In one case Q would assume the anti conformation (Q-T) and in the other the syn conformation (Q-A).…”

Section: Improving the Fidelity Of Tth Dna Ligasementioning

confidence: 99%

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Improving the Fidelity of Thermus Thermophilus DNA Ligase

Luo

Bergstrom

Barany

1996

Nucleic Acids Research

103

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The DNA ligase from Thermus thermophilus (Tth DNA ligase) seals single-strand breaks (nicks) in DNA duplex substrates. The specificity and thermostability of this enzyme are exploited in the ligase chain reaction (LCR) and ligase detection reaction (LDR) to distinguish single base mutations associated with genetic diseases. Herein, we describe a quantitative assay using fluorescently labeled substrates to study the fidelity of Tth DNA ligase. The enzyme exhibits significantly greater discrimination against all single base mismatches on the 3′-side of the nick in comparison with those on the 5′-side of the nick. Among all 12 possible single base pair mismatches on the 3′-side of the nick, only T-G and G-T mismatches generated a quantifiable level of ligation products after 23 h incubation. The high fidelity of Tth DNA ligase can be improved further by introducing a mismatched base or a universal nucleoside analog at the third position of the discriminating oligonucleotide. Finally, two mutant Tth DNA ligases, K294R and K294P, were found to have increased fidelity using this assay.

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Section: Synthesis Of Oligonucleotide Primersmentioning

confidence: 99%

Section: Improving the Fidelity Of Tth Dna Ligasementioning

confidence: 99%

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Improving the Fidelity of Thermus Thermophilus DNA Ligase

Luo

Bergstrom

Barany

1996

Nucleic Acids Research

103

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“…The efficiency of oligonucleotide primers containing multiple substitutions of 5-nitroindole (187, 188) and 3-nitropyrrole (189,190) has been studied and compared in DNA sequencing and in polymerase chain reactions. These analogs can be used at primer sites corresponding to degenerate base positions and specifically where the sequence data is incomplete.…”

Section: Universal Basesmentioning

confidence: 99%

MODIFIED OLIGONUCLEOTIDES: Synthesis and Strategy for Users

Verma

Eckstein²

1998

Annu. Rev. Biochem.

299

190

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Synthetic oligonucleotide analogs have greatly aided our understanding of several biochemical processes. Efficient solid-phase and enzyme-assisted synthetic methods and the availability of modified base analogs have added to the utility of such oligonucleotides. In this review, we discuss the applications of synthetic oligonucleotides that contain backbone, base, and sugar modifications to investigate the mechanism and stereochemical aspects of biochemical reactions. We also discuss interference mapping of nucleic acid-protein interactions; spectroscopic analysis of biochemical reactions and nucleic acid structures; and nucleic acid cross-linking studies.The automation of oligonucleotide synthesis, the development of versatile phosphoramidite reagents, and efficient scale-up have expanded the application of modified oligonucleotides to diverse areas of fundamental and applied biological research. Numerous reports have covered oligonucleotides for which modifications have been made of the phosphodiester backbone, of the purine and pyrimidine heterocyclic bases, and of the sugar moiety; these modifications serve as structural and mechanistic probes. In this chapter, we review the range, scope, and practical utility of such chemically modified oligonucleotides. Because of space limitations, we discuss only those oligonucleotides that contain phosphate and phosphate analogs as internucleotidic linkages. CONTENTS

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“…Moreover this type of compounds can give a deeper insight on the interactions with DNA since recently the 1-(2'-deoxy-β-D-ribofuranosyl)-3-nitropyrrole (2) was demonstrated to behave as universal nucleoside. 7 A unique property of this compound was its ability to replace long strings of contiguous nucleosides and still yield functional sequencing primers. Even if it does not resemble either a purine or pyrimidine nor does it contain a functional group of significant hydrogen bonding capacity, it showed an electronic distribution that resembled the average charge distribution of the DNA bases.…”

Section: Introductionmentioning

confidence: 99%