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Title: Double-stranded DNA analysis by matrix assisted laser desorption/ionization

Abstract

Matrix assisted laser desorption/ionization time-of-flight mass spectrometry was used to analyze double-stranded DNA fragments digested with restricted enzymes. As previously reported, only denatured single strands were detected and the resolution of TOFMS was not good enough to distinguish the mass difference between the complementary strands. To prove denaturation occurred rather than detecting doubly charged ds-DNA fragments, new DNA segments and restriction enzyme cleavage sites were searched. A 246 bp DNA designated as pLB129 was used. This segment contains part of the human immunoglobulin V{sub {lambda}} germline gene. It has several restriction sites that provide fragments with sticky ends. The 246 bp DNA was amplified by polyermase chain reaction (PCR), and digested with three kinds of restriction enzymes. The final products were purified with phenol/chloroform extraction and alcohol precipitation. MALDI-MS results clearly indicated that denaturation did happen as the mass peak corresponding to each individual was registered. 3-hydroxypicolinic acid (3-HPA) was used as the matrix. Analysis of double stranded DNA by MALDI-MS will have significant impacts in molecular biology. As coupled with PCR technique, MALDI-MS has great potential in fast screening tests for biomedical applications.

Authors:
 [1]; ; ;  [2];  [3]
  1. Vanderbilt Univ., Nashville, TN (United States)
  2. Oak Ridge National Lab., TN (United States)
  3. Univ. of Tennessee Medical Center, Knoxville, TN (United States)
Publication Date:
OSTI Identifier:
61957
Report Number(s):
CONF-9405234-
TRN: 95:003327-0092
DOE Contract Number:  
AC05-84OR21400
Resource Type:
Conference
Resource Relation:
Conference: 42. American Society of Mass Spectrometry (ASMS) conference on mass spectrometry and allied topics, Chicago, IL (United States), 29 May - 3 Jun 1994; Other Information: PBD: 1994; Related Information: Is Part Of Proceedings of the 42nd ASMS conference on mass spectrometry and allied topics; PB: 1233 p.
Country of Publication:
United States
Language:
English
Subject:
40 CHEMISTRY; 55 BIOLOGY AND MEDICINE, BASIC STUDIES; DNA; MASS SPECTROSCOPY; DESORPTION; TIME-OF-FLIGHT MASS SPECTROMETERS; SCREENING

Citation Formats

Tang, K, Taranenko, N I, Allman, S L, Chen, C H, and Chang, L Y. Double-stranded DNA analysis by matrix assisted laser desorption/ionization. United States: N. p., 1994. Web.
Tang, K, Taranenko, N I, Allman, S L, Chen, C H, & Chang, L Y. Double-stranded DNA analysis by matrix assisted laser desorption/ionization. United States.
Tang, K, Taranenko, N I, Allman, S L, Chen, C H, and Chang, L Y. Sat . "Double-stranded DNA analysis by matrix assisted laser desorption/ionization". United States.
@article{osti_61957,
title = {Double-stranded DNA analysis by matrix assisted laser desorption/ionization},
author = {Tang, K and Taranenko, N I and Allman, S L and Chen, C H and Chang, L Y},
abstractNote = {Matrix assisted laser desorption/ionization time-of-flight mass spectrometry was used to analyze double-stranded DNA fragments digested with restricted enzymes. As previously reported, only denatured single strands were detected and the resolution of TOFMS was not good enough to distinguish the mass difference between the complementary strands. To prove denaturation occurred rather than detecting doubly charged ds-DNA fragments, new DNA segments and restriction enzyme cleavage sites were searched. A 246 bp DNA designated as pLB129 was used. This segment contains part of the human immunoglobulin V{sub {lambda}} germline gene. It has several restriction sites that provide fragments with sticky ends. The 246 bp DNA was amplified by polyermase chain reaction (PCR), and digested with three kinds of restriction enzymes. The final products were purified with phenol/chloroform extraction and alcohol precipitation. MALDI-MS results clearly indicated that denaturation did happen as the mass peak corresponding to each individual was registered. 3-hydroxypicolinic acid (3-HPA) was used as the matrix. Analysis of double stranded DNA by MALDI-MS will have significant impacts in molecular biology. As coupled with PCR technique, MALDI-MS has great potential in fast screening tests for biomedical applications.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {1994},
month = {12}
}

Conference:
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