July 5
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| - | + | * Open Loop [Cc] | |
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| - | + | i) Cells were transferred to 50 mL Glu M9 in 250 mL flask with 5 ng/mL [aTc] and 1,0.1 uL/mL of inoculum. | |
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| + | ii) K12z1 was inoculated in 5 mL Glu M9 in 50 mL tube with [aTc]=5 ng/mL and 0.5,0.05 uL/mL of inoculum. | ||
| - | + | iii) However, right OD was not reached in time and this would be repeated. | |
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| - | + | * Open Loop [Dd] | |
| - | + | i)Cells were transferred to 50 mL Glu M9 in 250 mL flask with [aTc]=1 ng/mL and inoculum=1,0.1 uL/mL. | |
| - | + | ii) K12z1 was inoculated in 5 mL Glu M9 in 50 mL tube with [aTc]=1 ng/mL and inoculum=0.5,0.05 uL/mL. | |
| - | + | iii)The flask having cells with OD=0.193 was filtered out. | |
| - | + | iv) 7 dilutions with full IPTG range were made for pL.LuxR.Y.pR.C with AI from 1 ng/mL aTc and new 2xM9 in 1:1 ratio. | |
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| - | + | **-ve control: K12z1 in AI from 1 ng/mL aTc and new 2xM9 in 1:1 ratio | |
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| + | **+ve control: pL.Y, 100 uM IPTG, 1x M9 | ||
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| + | * To show that K12z1 does not produce any chemical in M9+aTc which might affect the system, we compared the CFP/YFP expressions of the following 2 samples: | ||
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| + | ** pL.LuxR.Y.pR.C was inoculated in Glu M9 with 500 uM IPTG, and | ||
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| + | ** K12Z1 was grown in 1 ng/mL aTc and the cells were filtered out. This medium was then diluted with 2xGlu M9 in a 1:1 ratio, and pL.LuxR.Y.pR.C was inoculated in it with 500 uM IPTG. | ||
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| + | * pL.LuxR.Y.pR.C, K12z1, pL.Y were Inoculated in LB | ||
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| + | * pL.LuxR.Y.pR.C and K12z1 were inoculated in LB. | ||
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| + | * pT.LuxI.C, K12Z1 were inoculated in LB for open loop duplicate expts [Ee] and [Cc]. | ||
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| + | '''Microscopy''' | ||
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| + | * Open loop samples [Aa] were imaged. | ||
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| + | * Open loop samples [Ff] were imaged. | ||
Revision as of 10:59, 6 July 2007
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- Open Loop [Cc]
i) Cells were transferred to 50 mL Glu M9 in 250 mL flask with 5 ng/mL [aTc] and 1,0.1 uL/mL of inoculum.
ii) K12z1 was inoculated in 5 mL Glu M9 in 50 mL tube with [aTc]=5 ng/mL and 0.5,0.05 uL/mL of inoculum.
iii) However, right OD was not reached in time and this would be repeated.
- Open Loop [Dd]
i)Cells were transferred to 50 mL Glu M9 in 250 mL flask with [aTc]=1 ng/mL and inoculum=1,0.1 uL/mL.
ii) K12z1 was inoculated in 5 mL Glu M9 in 50 mL tube with [aTc]=1 ng/mL and inoculum=0.5,0.05 uL/mL.
iii)The flask having cells with OD=0.193 was filtered out.
iv) 7 dilutions with full IPTG range were made for pL.LuxR.Y.pR.C with AI from 1 ng/mL aTc and new 2xM9 in 1:1 ratio.
- -ve control: K12z1 in AI from 1 ng/mL aTc and new 2xM9 in 1:1 ratio
- +ve control: pL.Y, 100 uM IPTG, 1x M9
- To show that K12z1 does not produce any chemical in M9+aTc which might affect the system, we compared the CFP/YFP expressions of the following 2 samples:
- pL.LuxR.Y.pR.C was inoculated in Glu M9 with 500 uM IPTG, and
- K12Z1 was grown in 1 ng/mL aTc and the cells were filtered out. This medium was then diluted with 2xGlu M9 in a 1:1 ratio, and pL.LuxR.Y.pR.C was inoculated in it with 500 uM IPTG.
- pL.LuxR.Y.pR.C, K12z1, pL.Y were Inoculated in LB
- pL.LuxR.Y.pR.C and K12z1 were inoculated in LB.
- pT.LuxI.C, K12Z1 were inoculated in LB for open loop duplicate expts [Ee] and [Cc].
Microscopy
- Open loop samples [Aa] were imaged.
- Open loop samples [Ff] were imaged.
