Difference between revisions of "Part:BBa K1628203"

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[[File:Pxyl_NK1.png]]
 
[[File:Pxyl_NK1.png]]
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We transformed the plasmids pHT01-''xylR'' and pCB-Pxyl into the NK-1 strain, to verify the activity of metabolic toggle switch (see it on our wiki). Fresh colonies of ''Bacillus amyloliquefaciens'' strains (NK-1 strain containing plasmids pHT01-''xylR'' and pCB-Pxyl and the control NK-1 strain containing plasmids pHT01 and pCB-Pxyl) were first cultured overnight in test tubes containing 5 mL LB liquid and then inoculated into 100 mL fresh fermentation medium. We added 1mM IPTG into the medium after 12h of cultivation. The β-galactosidase activity were measured at 12h, 18h, 24h, 30h, 36h, 42h to test the effect of metabolic toggle switch on the expression of ''bgaB''.
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As shown in Figure 2, β-galactosidase enzyme activity dropped considerably after 30 hours of fermentation. The inhibited expression of ''bgsB'' in experiment group (NPP+IPTG) indicated that the metabolic toggle switch we constructed is functional in ''B. amyloliquefaciens'' NK-1 strain.
  
 
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Revision as of 07:35, 18 September 2015

xylR

XylR is a repressor of xylose operon regulating promoter Pxyl. XylR together with promoter PlacI, repressor LacI (BBa_K1628201), promoter Pxyl (BBa_K1628002) and promoter Pxyl (BBa_K1628002) formed a metabolic toggle switch. We used this device to regulate the expression of odhAB genes in Bacillus amyloliquefaciens NK-1 (showed in Figure 2).

Pxyl NK1.png

We transformed the plasmids pHT01-xylR and pCB-Pxyl into the NK-1 strain, to verify the activity of metabolic toggle switch (see it on our wiki). Fresh colonies of Bacillus amyloliquefaciens strains (NK-1 strain containing plasmids pHT01-xylR and pCB-Pxyl and the control NK-1 strain containing plasmids pHT01 and pCB-Pxyl) were first cultured overnight in test tubes containing 5 mL LB liquid and then inoculated into 100 mL fresh fermentation medium. We added 1mM IPTG into the medium after 12h of cultivation. The β-galactosidase activity were measured at 12h, 18h, 24h, 30h, 36h, 42h to test the effect of metabolic toggle switch on the expression of bgaB.

As shown in Figure 2, β-galactosidase enzyme activity dropped considerably after 30 hours of fermentation. The inhibited expression of bgsB in experiment group (NPP+IPTG) indicated that the metabolic toggle switch we constructed is functional in B. amyloliquefaciens NK-1 strain.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 322
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]