Oligonucleotide sequencing using guanine-specific methylation and electrospray ionization ion trap mass spectrometry

Marzilli, Lisa A.; Barry, John P.; Sells, Todd B.; Law, Say-Jong; Vouros, Paul; Harsch, Andreas

doi:10.1002/(sici)1096-9888(199904)34:4<276::aid-jms757>3.0.co;2-o

Cited by 17 publications

(11 citation statements)

References 28 publications

(7 reference statements)

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“…48 Polyacrylamide gel electrophoresis is sensitive if an oligonucleotide is tagged with a radioactive or fluorescent label. There are some caveats about the use of the method.…”

Section: Purification and Characterization Of Modified Oligonucleotidesmentioning

confidence: 99%

“…The two most common methods used to evaluate purity of oligonucleotides are capillary gel electrophoresis and denaturing polyacrylamide gel electrophoresis (Scheme ). The former has very high revolving power with oligonucleotides and uses only trace accounts of material . Polyacrylamide gel electrophoresis is sensitive if an oligonucleotide is tagged with a radioactive or fluorescent label.…”

Section: 2 Purification and Characterization Of Modified Oligonucleot...mentioning

confidence: 99%

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Interactions of Carcinogen-Bound DNA with Individual DNA Polymerases

2006

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“…48 Polyacrylamide gel electrophoresis is sensitive if an oligonucleotide is tagged with a radioactive or fluorescent label. There are some caveats about the use of the method.…”

Section: Purification and Characterization Of Modified Oligonucleotidesmentioning

confidence: 99%

Section: 2 Purification and Characterization Of Modified Oligonucleot...mentioning

confidence: 99%

Interactions of Carcinogen-Bound DNA with Individual DNA Polymerases

2006

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“…Collision-induced dissociation (CID) of small oligonucleotides results in the formation of a-B and w ions as two major types of ions and thus can be used to sequence oligonucleotides (Scheme ) , . Recently, this approach (CID of oligonucleotide) has been used to detect normal APs, 3PG, N-7 alkylation, and other modifications in oligonucleotides − . Here, we present the use of ultraperformance liquid chromatography (UPLC)-coupled tandem MS-based approach for the detection of C4-AP at specific sites in DNA fragments without the need for hydrolysis of the DNA fragment or the use of specialized reagents.…”

Section: Introductionmentioning

confidence: 99%

Tandem Mass Spectrometry-Based Detection of C4′-Oxidized Abasic Sites at Specific Positions in DNA Fragments

Chowdhury

Guengerich

2009

Chem. Res. Toxicol.

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Oxidative damage to DNA has been linked to aging, cancer, and other biological processes. Reactive oxygen species and various antitumor agents including bleomycin and ionizing radiation have been shown to cause oxidative DNA sugar damage. Detection of DNA lesions is important for understanding the toxicological or therapeutic consequences associated with such agents. C4′-oxidized abasic sites (C4-AP) are produced by the antitumor drug bleomycin and ionizing radiation. The currently available methods for detection of C4-AP cannot provide both structural and sequence information. We have developed an LC- ESI-MS based approach for specific detection and mapping of C4-AP from a mixture of lesions. We show using Fe-bleomycin-damaged DNA, that C4-AP can be detected at cytosine and thymine sites by direct mass spectrometric (MS) analysis. Our results reveal that collision-induced dissociation of C4-AP-containing oligonucleotides results in preferential fragmentation at C4-AP sites with the formation of the unique a* ions (18 a.m.u more than the a-B ions) that allow mapping of the C4-AP sites. Various chemical modification strategies (e.g. reduction with NaBH4 and NaBD4 and derivatization with methoxyamine and hydrazine, followed by liquid-chromatography (LC)-MS analysis) were also used for unambiguous detection of C4-AP sites. Finally, we show that the methods described here can detect the presence of C4-AP at specific sites in a complex sample such as hydroxyl radical-damaged DNA. The LC-MS approach was also used for the simultaneous detection of the other C4′-oxidation end product, 3′-phosphoglycolate (3PG) at a specific site in hydroxyl radical-damaged DNA. Thus, LCMS provides a rapid and direct approach for the detection and mapping of oxidative DNA lesions.

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“…Traditional collision-based fragmentation, i.e., collision-activated dissociation via resonant excitation (CAD) and beam-type CAD (a.k.a., HCD), are among the most studied tandem MS strategies because of their accessibility and ease of application to deprotonated precursors . For RNA anion dissociation, these methods direct cleavage across numerous channels, often leading to highly complex spectra and the generation of internal fragments and neutral losses (which can include the loss of RNA nucleobases themselves). ,, McLuckey and others demonstrated the dependence of charge-state and activation energy on product ion distributions and, for CAD, these drawbacks can be reduced for precursors having low charge states by minimizing collisional energy. − …”

mentioning

confidence: 99%

“…21 For RNA anion dissociation, these methods direct cleavage across numerous channels, often leading to highly complex spectra and the generation of internal fragments and neutral losses (which can include the loss of RNA nucleobases themselves). 11,22,23 McLuckey and others demonstrated the dependence of charge-state and activation energy on product ion distributions and, for CAD, these drawbacks can be reduced for precursors having low charge states by minimizing collisional energy. 24−26 The ideal method for dissociating RNA anions would be indifferent to precursor length, charge, and the presence of modifications.…”

mentioning

confidence: 99%

Ribonucleic Acid Sequence Characterization by Negative Electron Transfer Dissociation Mass Spectrometry

et al. 2020

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Modified oligonucleotides represent a promising avenue for drug development, with small interfering RNAs (siRNA) and microRNAs gaining traction in the therapeutic market. Mass spectrometry (MS)-based analysis offers many benefits for characterizing modified nucleic acids. Negative electron transfer dissociation (NETD) has proven valuable in sequencing oligonucleotide anions, particularly because it can retain modifications while generating sequence-informative fragments. We show that NETD can be successfully implemented on a widely available quadrupole-Orbitrap-linear ion trap mass spectrometer that uses a front-end glow discharge source to generate radical fluoranthene reagent cations. We characterize both unmodified and modified ribonucleic acids and present the first application of activated-ion negative electron transfer dissociation (AI-NETD) to nucleic acids. AI-NETD achieved 100% sequence coverage for both a 6-mer (5′-rGmUrArCmUrG-3′) with 2′-O-methyl modifications and a 21-mer (5′-rCrArUrCrCrUrCrUrArGrArGrGrArUrArGrArArUrG-3′), the luciferase antisense siRNA. Both NETD and AI-NETD afforded complete sequence coverage of these molecules while maintaining a relatively low degree of undesired base-loss products and internal products relative to collision-based methods.

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Oligonucleotide sequencing using guanine-specific methylation and electrospray ionization ion trap mass spectrometry

Cited by 17 publications

References 28 publications

Interactions of Carcinogen-Bound DNA with Individual DNA Polymerases

Interactions of Carcinogen-Bound DNA with Individual DNA Polymerases

Tandem Mass Spectrometry-Based Detection of C4′-Oxidized Abasic Sites at Specific Positions in DNA Fragments

Ribonucleic Acid Sequence Characterization by Negative Electron Transfer Dissociation Mass Spectrometry

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