Difference between revisions of "Part:BBa K1628202"

Revision as of 07:34, 18 September 2015

Pgrac

Promoter Pgrac is a promoter of lactose operon regulated by repressor LacI. Pgrac together with promoter PlacI, repressor LacI (BBa_K1628201), promoter Pxyl (BBa_K1628002) and repressor XylR (BBa_K1628203) 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).

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
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

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 [[File: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.
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