Difference between revisions of "Part:BBa K3861003"
 
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<i>hilA<\i>, together with <i>hilC<\i> and <i>hilD<\i>, are global regulators and control the expression of the  <i>Salmonella<\i> pathogenicity island I(SPI-1) genes in <i>S<\i>. typhimurium on which the SPI-1 T3SS injectisome is encoded (Widmaier et al., 2009). Over 25 genes that are needed for host invasion are located on the SPI-1 (Darwin and Miller, 1999). Also, <i>hilA<\i> activates SPI-1 operon mediated by the prg and inv promoters (Bajaj and Lee et al., 1995; Bajaj et al., 1996).<i>hilA<\i> controls <i>SPI<\i> genes in response to enviromental needs like pH and oxygen levels (Ahmer et al., 1999; Bajaj et al., 1996, Main-Hester et al., 2008).
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The coding region of the Thymidylate synthase (ThyA, EC2.1.1.45). The enzyme catalyzes the reductive methylation of 2'-deoxyuridine-5'-monophosphate (dUMP) to 2'-deoxythymidine-5'-monophosphate (dTMP). <i>Salmonella</i> Typhimurium <i>thyA</i> knock out mutants cannot synthesize DNA if no thymidine monophosphate (dTMP) is available to the cells.<sup>1</sup> Furthermore, <i>thyA</i> mutants were shown to not grow intercellular in macrophage-like and  human epithelial cell lines <i>in vitro</i>.<sup>2</sup> Thus, for dTMP auxotrophic bacterial strains can be created. We used this knowledge to our advantage and used <i>thyA</i> as selection marker. By deleting the chromosomal <i>thyA</i> gene and reintroducing it with one of our plasmids (<a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K3861005">BBa_K3861005</a>), we effectively bypassed the need for an antibiotic based selection marker system.    
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<partinfo>BBa_K3861003 parameters</partinfo>
 
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==='''References'''===
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1.      Neuhard, J. & Kelln, R. A. (1996). Biosynthesis and conversions ofpyrimidines. In Escherichia coli and Salmonella, pp. 580–599. <i>American Society for Microbiology</i>
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2. Kok, M., Bühlmann, E. & Pechère, J.-C. 2001. Salmonella typhimurium thyA mutants fail to grow intracellularly in vitro and are attenuated in mice. <i>Microbiology 147</i>, 727–733.

Latest revision as of 15:55, 21 October 2021


ThyA CDS

The coding region of the Thymidylate synthase (ThyA, EC2.1.1.45). The enzyme catalyzes the reductive methylation of 2'-deoxyuridine-5'-monophosphate (dUMP) to 2'-deoxythymidine-5'-monophosphate (dTMP). Salmonella Typhimurium thyA knock out mutants cannot synthesize DNA if no thymidine monophosphate (dTMP) is available to the cells.1 Furthermore, thyA mutants were shown to not grow intercellular in macrophage-like and human epithelial cell lines in vitro.2 Thus, for dTMP auxotrophic bacterial strains can be created. We used this knowledge to our advantage and used thyA as selection marker. By deleting the chromosomal thyA gene and reintroducing it with one of our plasmids (BBa_K3861005), we effectively bypassed the need for an antibiotic based selection marker system.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


References

1. Neuhard, J. & Kelln, R. A. (1996). Biosynthesis and conversions ofpyrimidines. In Escherichia coli and Salmonella, pp. 580–599. American Society for Microbiology
2. Kok, M., Bühlmann, E. & Pechère, J.-C. 2001. Salmonella typhimurium thyA mutants fail to grow intracellularly in vitro and are attenuated in mice. Microbiology 147, 727–733.