Rickettsial ompB Promoter Regulated Expression of GFPuv in Transformed Rickettsia montanensis

被引:14
作者
Baldridge, Gerald D. [1 ]
Burkhardt, Nicole Y. [1 ]
Oliva, Adela S. [1 ]
Kurtti, Timothy J. [1 ]
Munderloh, Ulrike G. [1 ]
机构
[1] Univ Minnesota, Dept Entomol, St Paul, MN 55108 USA
来源
PLOS ONE | 2010年 / 5卷 / 01期
关键词
GREEN FLUORESCENT PROTEIN; FEVER GROUP RICKETTSIAE; MOUNTAIN WOOD TICK; SPOTTED-FEVER; DERMACENTOR-ANDERSONI; IXODES-SCAPULARIS; TRANSPOSON MUTAGENESIS; WESTERN MONTANA; GENE-EXPRESSION; CAPE-COD;
D O I
10.1371/journal.pone.0008965
中图分类号
O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];
学科分类号
07 ; 0710 ; 09 ;
摘要
Background: Rickettsia spp. (Rickettsiales: Rickettsiaceae) are Gram-negative, obligate intracellular, alpha-proteobacteria that have historically been associated with blood-feeding arthropods. Certain species cause typhus and spotted fevers in humans, but others are of uncertain pathogenicity or may be strict arthropod endosymbionts. Genetic manipulation of rickettsiae should facilitate a better understanding of their interactions with hosts. Methodology/Principal Findings: We transformed a species never associated with human disease, Rickettsia montanensis, by electroporation with a TN5 transposon (pMOD700) containing green fluorescent protein (GFPuv) and chloramphenicol acetyltransferase (CAT) genes under regulation of promoters cloned from the Rickettsia rickettsii ompB gene, and isolated a Chloramphenicol-resistant GFP-fluorescent rickettsiae population (Rmontanensis700). The Rmontanensis700 rickettsiae contained a single transposon integrated near an acetyl-CoA acetyltransferase gene in the rickettsial chromosome. Northern blots showed that GFPuv and CAT mRNAs were both expressed as two transcripts of larger and smaller than predicted length. Western immunoblots showed that Rmontanensis700 and E. coli transformed with a plasmid containing the pMOD700 transposon both expressed GFPuv proteins of the predicted molecular weight. Conclusions/Significance: Long-standing barriers to transformation of rickettsiae have been overcome by development of transposon-based rickettsial transformation vectors. The ompB promoter may be the most problematic of the four promoters so far employed in those vectors.
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页数:8
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共 52 条
  • [1] Spotted-fever group Rickettsia in Dermacentor variabilis, Maryland
    Ammerman, NC
    Swanson, KI
    Anderson, JM
    Schwartz, TR
    Seaberg, EC
    Glass, GE
    Norris, DE
    [J]. EMERGING INFECTIOUS DISEASES, 2004, 10 (08) : 1478 - 1481
  • [2] A PROTECTIVE PROTEIN ANTIGEN OF RICKETTSIA-RICKETTSII HAS TANDEMLY REPEATED, NEAR-IDENTICAL SEQUENCES
    ANDERSON, BE
    MCDONALD, GA
    JONES, DC
    REGNERY, RL
    [J]. INFECTION AND IMMUNITY, 1990, 58 (09) : 2760 - 2769
  • [3] RICKETTSIA-RICKETTSII AND RICKETTSIA-MONTANA FROM IXODID TICKS IN CONNECTICUT
    ANDERSON, JF
    MAGNARELLI, LA
    PHILIP, RN
    BURGDORFER, W
    [J]. AMERICAN JOURNAL OF TROPICAL MEDICINE AND HYGIENE, 1986, 35 (01) : 187 - 191
  • [4] Rickettsial pathogens and their arthropod vectors
    Azad, AF
    Beard, CB
    [J]. EMERGING INFECTIOUS DISEASES, 1998, 4 (02) : 179 - 186
  • [5] Sequence and expression analysis of the ompA gene of Rickettsia peacockii, an endosymbiont of the rocky mountain wood tick, Dermacentor andersoni
    Baldridge, GD
    Burkhardt, NY
    Simser, JA
    Kurtti, TJ
    Munderloh, UG
    [J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2004, 70 (11) : 6628 - 6636
  • [6] Analysis of fluorescent protein expression in transformants of Rickettsia monacensis, an obligate intracellular tick symbiont
    Baldridge, GD
    Burkhardt, N
    Herron, MJ
    Kurtti, TJ
    Munderloh, UG
    [J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2005, 71 (04) : 2095 - 2105
  • [7] Plasmids of the pRM/pRF family occur in diverse Rickettsia species
    Baldridge, Gerald D.
    Burkhardt, Nicole Y.
    Felsheim, Roderick F.
    Kurtti, Timothy J.
    Munderloh, Ulrike G.
    [J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2008, 74 (03) : 645 - 652
  • [8] Transposon insertion reveals pRM, a plasmid of Rickettsia monacensis
    Baldridge, Gerald D.
    Burkhardt, Nicole Y.
    Felsheim, Roderick F.
    Kurtti, Timothy J.
    Munderloh, Ulrike G.
    [J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2007, 73 (15) : 4984 - 4995
  • [9] Infection of Ixodes scapularis ticks with Rickettsia monacensis expressing green fluorescent protein:: A model system
    Baldridge, Gerald D.
    Kurtti, Timothy J.
    Burkhardt, Nicole
    Baldridge, Abigail S.
    Nelson, Curtis M.
    Oliva, Adela S.
    Munderloh, Ulrike G.
    [J]. JOURNAL OF INVERTEBRATE PATHOLOGY, 2007, 94 (03) : 163 - 174
  • [10] BELL EJ, 1963, J IMMUNOL, V90, P770