Monitoring viral DNA release with capillary electrophoresis

被引:6
作者
Krylova, SM
Rozenberg, D
Coulton, JW
Krylov, SN [1 ]
机构
[1] York Univ, Dept Chem, N York, ON M3J 1P3, Canada
[2] McGill Univ, Dept Microbiol & Immunol, Montreal, PQ H3A 2B4, Canada
关键词
D O I
10.1039/b409835h
中图分类号
O65 [分析化学];
学科分类号
070302 ; 081704 ;
摘要
Viral DNA injection into host cells is one of the primary mechanisms of viral propagation. Drug development that targets viral propagation requires fast and sensitive methods for monitoring the release of viral DNA in vitro. Here we demonstrate the use of capillary electrophoresis (CE) for monitoring DNA release from virus particles. As a model for this study, we used T5 bacteriophages that infect the bacterium Escherichia coli K-12 by binding to the outer membrane FhuA receptor and then injecting DNA. DNA release from the T5 phages in vitro was induced by either elevated temperature or by interaction with the purified FhuA receptor. After DNA release, the viral samples were stained with the high affinity fluorescent dye YOYO-1, injected into the capillary and subjected to electrophoresis. YOYO-1-stained DNA generated a well-defined peak, allowing reliable detection of viral DNA from as few as 10(5) viral particles. The staining to track T5 phage DNA release exemplifies the great versatility that CE offers in studying viral systems. This CE-based method can be used to study molecular mechanisms of viral infections and to evaluate anti-viral drug candidates.
引用
收藏
页码:1234 / 1237
页数:4
相关论文
共 14 条
  • [1] Factors affecting M13 bacteriophage inactivation by methylene blue photosensitization
    Abe, H
    Wagner, SJ
    Kuwabara, M
    Kamo, N
    Ikebuchi, K
    Sekiguchi, S
    [J]. PHOTOCHEMISTRY AND PHOTOBIOLOGY, 1997, 65 (05) : 873 - 876
  • [2] Purification and structural and functional characterization of FhuA, a transporter of the Escherichia coli outer membrane
    Boulanger, P
    leMaire, M
    Bonhivers, M
    Dubois, S
    Desmadril, M
    Letellier, L
    [J]. BIOCHEMISTRY, 1996, 35 (45) : 14216 - 14224
  • [3] Flow cytometric detection of viruses
    Brussaard, CPD
    Marie, D
    Bratbak, G
    [J]. JOURNAL OF VIROLOGICAL METHODS, 2000, 85 (1-2) : 175 - 182
  • [4] Ferguson AD, 1998, PROTEIN SCI, V7, P1636, DOI 10.1002/pro.5560070719
  • [5] Siderophore-mediated iron transport: Crystal structure of FhuA with bound lipopolysaccharide
    Ferguson, AD
    Hofmann, E
    Coulton, JW
    Diederichs, K
    Welte, W
    [J]. SCIENCE, 1998, 282 (5397) : 2215 - 2220
  • [6] FUNCTIONAL INTERACTION OF TONA-TONB RECEPTOR SYSTEM IN ESCHERICHIA-COLI
    HANTKE, K
    BRAUN, V
    [J]. JOURNAL OF BACTERIOLOGY, 1978, 135 (01) : 190 - 197
  • [7] FLUORESCENTLY LABELED VIRUS PROBES SHOW THAT NATURAL VIRUS POPULATIONS CAN CONTROL THE STRUCTURE OF MARINE MICROBIAL COMMUNITIES
    HENNES, KP
    SUTTLE, CA
    CHAN, AM
    [J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 1995, 61 (10) : 3623 - 3627
  • [8] Binding of fluorescent dye to genomic RNA inside intact human rhinovirus after viral capsid penetration investigated by capillary electrophoresis
    Kremser, L
    Okun, VM
    Nicodemou, A
    Blaas, D
    Kenndler, E
    [J]. ANALYTICAL CHEMISTRY, 2004, 76 (04) : 882 - 887
  • [9] Marie D, 1999, APPL ENVIRON MICROB, V65, P45
  • [10] The thermodynamic basis of mechanisms of bacterial virus infection
    Mrevlishvili, GM
    Mdzinarashvili, T
    Al-Zaza, M
    Tsinadze, L
    Tushishvili, D
    Razmadze, G
    [J]. PURE AND APPLIED CHEMISTRY, 1999, 71 (07) : 1291 - 1299