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Microbial Cell Biology

Formation of dTDP-Rhamnose Is Essential for Growth of Mycobacteria

Yufang Ma, Fei Pan, Michael McNeil
Yufang Ma
Department of Microbiology, Colorado State University, Fort Collins, Colorado 80523
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Fei Pan
Department of Microbiology, Colorado State University, Fort Collins, Colorado 80523
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Michael McNeil
Department of Microbiology, Colorado State University, Fort Collins, Colorado 80523
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  • For correspondence: mmcneil@colostate.edu
DOI: 10.1128/JB.184.12.3392-3395.2002
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    FIG. 1.

    Southern blot analysis of M. smegmatis. (A-1) Southern blot of DNA from the first-crossover strain, M. smegmatis FP201 (Table 1). DNA was cleaved with SmaI, and the probe was rmlD. (A-2) Origins of the fragments. (B-1) Southern blot of DNA from the second-crossover strain, M. smegmatis YM202 (Table 1). DNA was cleaved with SmaI, and the probe was rmlD. (B-2) Origins of the fragments. In the original blot, a fragment at 10 kb that originated from the rescue plasmid was faintly visible. Values to the sides of the blots are molecular sizes (in kilobases).

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    FIG. 2.

    Growth curves of M. smegmatis strains at 30 and 42°C. The growth of the control strain, M. smegmatis mc2155, containing plasmid pYM201 at 30°C (▴) and at 42°C (▵), and the growth of the rmlD knockout strain, M. smegmatis YM202, containing plasmid pYM201 at 30°C (•) and at 42°C (○), are illustrated.

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  • TABLE 1.

    Key bacterial strains and plasmids

    Strain or plasmidRelevant characteristic(s)Reference or source
    Strains
        M. smegmatis mc2 155Harbors all plasmids used herein 22
        M. smegmatis FP201 M. smegmatis mc2 155 with pFP201 integrated into the rmlD locus (Fig. 1)This study
        M. smegmatis YM202 M. smegmatis FP201 which has undergone a second-crossover event in the presence of pYM201 (Fig. 1)This study
    Plasmids
        pPR27TS mycobacterial origin of replication; carries sacB gene, gen, E. coli origin of replication 21
        pFP201pPR27 derivative carrying rmlD::kan and the xylE geneThis study
        pCG76 Escherichia coli/Mycobacterium shuttle vector carrying a TS mycobacterial origin of replication and streptomycin resistance cassette (str) 11
        pYM201TS rescue plasmid pCG76 carrying the M. tuberculosis rmlD gene under the control of the rmlD natural promoter, strThis study
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Formation of dTDP-Rhamnose Is Essential for Growth of Mycobacteria
Yufang Ma, Fei Pan, Michael McNeil
Journal of Bacteriology Jun 2002, 184 (12) 3392-3395; DOI: 10.1128/JB.184.12.3392-3395.2002

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Formation of dTDP-Rhamnose Is Essential for Growth of Mycobacteria
Yufang Ma, Fei Pan, Michael McNeil
Journal of Bacteriology Jun 2002, 184 (12) 3392-3395; DOI: 10.1128/JB.184.12.3392-3395.2002
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    • ABSTRACT
    • Constructing the rmlD replacement plasmid (pFP201) and obtaining the first homologous recombination event.
    • Construction of an rmlD rescue plasmid.
    • Second-crossover attempts and events.
    • M. smegmatis YM202 will not grow at 42°C.
    • The wbbL gene is transcribed and translated in YM202.
    • Application of results to M. tuberculosis.
    • ACKNOWLEDGMENTS
    • FOOTNOTES
    • REFERENCES
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KEYWORDS

Mycobacterium smegmatis
Nucleoside Diphosphate Sugars
Thymine Nucleotides

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