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Team:Chiba/lux promoter
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==Abstract== | ==Abstract== | ||
| - | LuxR is an AHL-dependent activator. However | + | LuxR is an AHL-dependent activator.LuxR-AHL complex bind lux box,a 20-bp sequence and activate transcription. However the lux box is inserted between -35 and -10 and LuxR functions as LuxR-AHL inverter (Plux inv) .The Plux inv was resistered in Biobrick number R0061.We've constructed Plux inv-GFP combinig R0061and E0240 and characterized about it.<br> |
| - | + | ||
==Experiments== | ==Experiments== | ||
===1.Preparing Plux inv-GFP=== | ===1.Preparing Plux inv-GFP=== | ||
| + | Plasmid:R0061 and E0240<br> | ||
| + | Strain:XL10-G<br> | ||
| + | protocol<br> | ||
Preparing process of Plux inv-GFP is shown in '''Fig. 1'''.We transformed preparing Plux inv-GFP in XL10-G and confirmed the expression GFP Fluorescence.The sequence is also conformed. | Preparing process of Plux inv-GFP is shown in '''Fig. 1'''.We transformed preparing Plux inv-GFP in XL10-G and confirmed the expression GFP Fluorescence.The sequence is also conformed. | ||
| Line 163: | Line 165: | ||
===2.Characterization Plux inv-GFP=== | ===2.Characterization Plux inv-GFP=== | ||
| - | Plux inv-GFP | + | |
| - | + | Plasmid:Plux inv-GFP and Plac-LuxR (LuxR constitutive generator)<br> | |
| - | We cotransformded Plux inv-GFP and LuxR | + | strain:DH10B<br> |
| - | + | ||
| + | protocol<br> | ||
| + | We cotransformded Plux inv-GFP and LuxR Plac-LuxR in DH10B. | ||
| + | 1,LBL | ||
| + | incubation for12 h at 37゜C. | ||
==Results== | ==Results== | ||
| - | + | ||
| - | + | We confirmed GFP Fluorescence both AHL+ and -sample.<br> | |
| - | [[Image:Plux inv results.png|frame| | + | <br> |
| + | [[Image:Plux inv results.png|frame|none|Fig. 3 ]] | ||
| + | |||
| + | ==discussion== | ||
| + | 私たちの系では，LuxRの抑制を確認することができなかった。 | ||
==Reference== | ==Reference== | ||
#Egland.K.A, and Greenberg.E.P, Conversion of the ''Vibrio Fischeri'' Transcriptional Activator,LuxR, to a Repressor, ''J. Bacteriol.'', '''182''', P.805-811 (2000) | #Egland.K.A, and Greenberg.E.P, Conversion of the ''Vibrio Fischeri'' Transcriptional Activator,LuxR, to a Repressor, ''J. Bacteriol.'', '''182''', P.805-811 (2000) | ||
#Cox.R.S.3rd, Surette.M.G, Elowitz.M.B, Programming gene expression with combinatorial promoters, ''Mol Syst Biol'', '''3''' ,145 (2007) | #Cox.R.S.3rd, Surette.M.G, Elowitz.M.B, Programming gene expression with combinatorial promoters, ''Mol Syst Biol'', '''3''' ,145 (2007) | ||
Latest revision as of 13:29, 24 January 2011
Contents |
Abstract
LuxR is an AHL-dependent activator.LuxR-AHL complex bind lux box,a 20-bp sequence and activate transcription. However the lux box is inserted between -35 and -10 and LuxR functions as LuxR-AHL inverter (Plux inv) .The Plux inv was resistered in Biobrick number R0061.We've constructed Plux inv-GFP combinig R0061and E0240 and characterized about it.
Experiments
1.Preparing Plux inv-GFP
Plasmid:R0061 and E0240
Strain:XL10-G
protocol
Preparing process of Plux inv-GFP is shown in Fig. 1.We transformed preparing Plux inv-GFP in XL10-G and confirmed the expression GFP Fluorescence.The sequence is also conformed.
2.Characterization Plux inv-GFP
Plasmid:Plux inv-GFP and Plac-LuxR (LuxR constitutive generator)
strain:DH10B
protocol
We cotransformded Plux inv-GFP and LuxR Plac-LuxR in DH10B.
1,LBL
incubation for12 h at 37゜C.
Results
We confirmed GFP Fluorescence both AHL+ and -sample.
discussion
私たちの系では，LuxRの抑制を確認することができなかった。
Reference
- Egland.K.A, and Greenberg.E.P, Conversion of the Vibrio Fischeri Transcriptional Activator,LuxR, to a Repressor, J. Bacteriol., 182, P.805-811 (2000)
- Cox.R.S.3rd, Surette.M.G, Elowitz.M.B, Programming gene expression with combinatorial promoters, Mol Syst Biol, 3 ,145 (2007)
