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Food Microbiology

Identification of Listeria monocytogenes Genes Expressed in Response to Growth at Low Temperature

Siqing Liu, James E. Graham, Lance Bigelow, Philip D. Morse, II, Brian J. Wilkinson
Siqing Liu
Microbiology Group, Department of Biological Sciences
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James E. Graham
Division of Infectious Diseases, Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee 37232
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Lance Bigelow
Microbiology Group, Department of Biological Sciences
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Philip D. Morse
Department of Chemistry, Illinois State University, Normal, Illinois 61790
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Brian J. Wilkinson
Microbiology Group, Department of Biological Sciences
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  • For correspondence: bjwilkin@ilstu.edu
DOI: 10.1128/AEM.68.4.1697-1705.2002
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  • FIG. 1.
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    FIG. 1.

    Schematic diagram outlining the procedure (SCOTS) used to obtain cDNAs for L. monocytogenes RNAs expressed in response to growth at a low temperature. (A) Process used to reduce rDNA sequence content and increase the representation of less abundant RNAs (normalized). (B) Differential hybridization method used for selective amplification of bacterial RNAs expressed in response to a reduced temperature. S:B, streptavidin:biotin.

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    FIG. 2.

    Northern hybridization analyses of RNAs from mid-log-phase L. monocytogenes 10403S cultures grown at 37 and 10°C. EcoRI-digested inserts from plasmid cDNA clones obtained by SCOTS were used as templates to prepare radiolabeled probes. Hybridization was performed as described in Materials and Methods. Nylon filters were subsequently hybridized with a 16S rDNA probe and a housekeeping gene (dapE) probe as controls. Table 2 lists the corresponding genes for SCOTS cDNA clones.

  • FIG. 3.
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    FIG. 3.

    Northern hybridization analyses of L. monocytogenes RNAs. Total RNA was isolated from mid-log-phase L. monocytogenes 10403S grown at either 37 or 10°C. RNAs were electrophoresed as described in Materials and Methods and stained with ethidium bromide to ensure that equivalent amounts of total RNA were present in the lanes (bottom panel). EcoRI-digested inserts from plasmid cDNA clones obtained by SCOTS were used as templates to prepare radiolabeled probes (top panel). Table 2 lists the genes corresponding to SCOTS cDNA clones.

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  • TABLE 1.

    Oligonucleotides used for this study

    OligonucleotideSequence
    F1CTGGCTCAGGACGAACGCTGG
    B1GCTGGCTCCATAAAGGTGACC
    F2TCGGTAGGGTCACCTTTATGG
    B2CTCACACTCACTGCTTGGACG
    F3AATGGAAATGTGCGTCCAAGC
    B3ACTTTTCAAATTGCCCGGCAG
    K9GACACTCTCGAGACATCACCGGTACCNNNNNN
    F9GCCGGAGCTCTGCAGAATTCNNNNNN
    K-N6GACACTCTCGAGACATCACCGGTACC
    F-N6GCCGGAGCTCTGCAGAATTC
    DapE5′GGTTTTGGCATTTTCAGGGC
    DapE3′GATTCTGTGTCCACTCGGTTCG
  • TABLE 2.

    L. monocytogenes cDNA clones identified by SCOTS

    CategorySCOTS clone(s)GenePredicted gene productGenBank accession no.Strain EGD-e
    GeneChromosomal location (kb)
    Previously described L. monocytogenes cold-adaptive responses12flaAFlagellinX65624lmo0690724.9
    25flpFerritin-like proteinAJ244014lmo0943979.1
    Regulatory adaptive responses19, 62rpoN (sigL)RNA polymerase sigma factor σ54X93169.1lmo24612535.2
    73lhkAHistidine kinase sensorAJ010495lmo15081538.5
    6yycJTwo-component signal transduction systemBAA11296lmo0291316.9
    22bglGSimilar to transcription antiterminator BglG familylmo0501536.3
    75adaBSimilar to methyltransferaseX53399lmo0571608.5
    General microbial stress responses70groELMajor heat shock proteinAF335323lmo20682149.3
    55clpPSerine proteaseAF102775lmo26122627.7
    7trxBThioredoxin reductaseAF009622lmo24782554.4
    60clpBClpB proteaselmo22062297.2
    Alterations in amino acid metabolism17, 24, 45hisJHistidinol phosphate phosphataseD70002lmo0570607.4
    18trpGAnthranilate synthaseS74362lmo27492824.1
    32cysSCysteinyl-tRNA synthetaseL14580lmo0239259.5
    65aroA3-Deoxy-d-arabino-heptulosonate 7-phosphate synthaseX69545lmo16001645.2
    51celDEndoglucanase DU07818lmo1719178.1
    Alterations in degradative metabolism15, 27, 39, 43, 64, 84eutBEthanolamine ammonia lyase heavy chainP19635lmo11751203.6
    79mleASimilar to malolactic enzymeX82326lmo19151988.2
    Alterations of the L. monocytogenes cell surface3psrPBP5 synthesis repressorU42211lmo0443472.8
    56fbpFibronectin-binding proteinAJ132543lmo0721751.3
    Proteins with unknown functions4Unknown conserved hypothetical proteinlmo0599lmo0599640.3
    26Unknown proteinlmo0170lmo0170168.0
    50Unknown proteinlmo0719lmo0719750.0
    69Unknown proteinlmo1535lmo15351570.9
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Identification of Listeria monocytogenes Genes Expressed in Response to Growth at Low Temperature
Siqing Liu, James E. Graham, Lance Bigelow, Philip D. Morse II, Brian J. Wilkinson
Applied and Environmental Microbiology Apr 2002, 68 (4) 1697-1705; DOI: 10.1128/AEM.68.4.1697-1705.2002

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Identification of Listeria monocytogenes Genes Expressed in Response to Growth at Low Temperature
Siqing Liu, James E. Graham, Lance Bigelow, Philip D. Morse II, Brian J. Wilkinson
Applied and Environmental Microbiology Apr 2002, 68 (4) 1697-1705; DOI: 10.1128/AEM.68.4.1697-1705.2002
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