Test Mnt/Tet promoter - Revision history

BoCool at 16:43, 4 August 2008

2008-08-04T16:43:37Z

BoCool at 14:24, 18 July 2008

2008-07-18T14:24:19Z

BoCool at 16:27, 15 July 2008

2008-07-15T16:27:25Z

BoCool at 15:31, 3 July 2008

2008-07-03T15:31:29Z

BoCool at 15:01, 3 July 2008

2008-07-03T15:01:16Z

BoCool at 19:53, 2 July 2008

2008-07-02T19:53:34Z

BoCool at 19:49, 2 July 2008

2008-07-02T19:49:40Z

BoCool at 15:18, 2 July 2008

2008-07-02T15:18:18Z

BoCool at 14:42, 2 July 2008

2008-07-02T14:42:12Z

BoCool at 14:28, 2 July 2008

2008-07-02T14:28:52Z

@@ Line 83: / Line 83: @@
 This makes the third promoter test sequence
 the first and third test sequences will also need to be tested by adding aTc
 the first test sequence should glow and the third should not.

@@ Line 25: / Line 25: @@
 I13453+[http://partsregistry.org/Part:BBa_S0101 S0101]+[http://partsregistry.org/Part:BBa_B0015 B0015]+[http://partsregistry.org/Part:BBa_K091105 K091105]+[http://partsregistry.org/Part:BBa_E0240 E0240]
 add aTc
@@ Line 38: / Line 42: @@
 I13453+[http://partsregistry.org/Part:BBa_I13025 I13025]+B0015+K091105+E0240

@@ Line 53: / Line 53: @@
 ) K091105 (SP) ligated with E0240 (XP)
-) need to ligate pBAD (RS) to and RBS (RX)
+) ligate pBAD+RBS+TetR+TT (from build XOR gate page) to the 5' end of pMnt/Tet+RBS+GFP
 This will make the first promoter test sequence.

@@ Line 51: / Line 51: @@
 '''Ligation plan for three test sequences'''
-) K091105 ligated with E0240
+) K091105 (SP) ligated with E0240 (XP)
-) need to ligate pBAD to and RBS
+) need to ligate pBAD (RS) to and RBS (RX)
-) then ligate TetR to the 3' end of that
+) then ligate TetR (XP) to the 3' end of that (SP)
-)then ligate TT to the 3' end of that
+)then ligate TT (RX) to the 3' end of that (RS)
-)then ligate that part 5' to the pMnt/Tet+GFP
+)then ligate that part (RX) 5' to the pMnt/Tet+GFP (RS)
 This will make the first promoter test sequence.
@@ Line 65: / Line 65: @@
-) ligate the part RBS+Mnt to the 5' end of the TT
+) ligate the part RBS+Mnt (RS) to the 5' end of the TT (RX)
-) then ligate pBAD to the 5' end of that
+) then ligate pBAD (RS) to the 5' end of that (RX)
-) then ligate the part pMnt/Tet+RBS+GFP to the 3' end of that
+) then ligate the part pMnt/Tet+RBS+GFP (RS) to the 3' end of that (RX)
 This makes the second promoter test sequence.
@@ Line 75: / Line 75: @@
-) ligate the part RBS+Mnt+TT to the 3' end of pBAD+RBS+TetR
+) ligate the part RBS+Mnt+TT (RX) to the 3' end of pBAD+RBS+TetR (RS)
-) then ligate the part pMnt/Tet+RBS+GFP to the 3' end of that
+) then ligate the part pMnt/Tet+RBS+GFP (SP) to the 3' end of that (XP)
-) This makes the third promoter test sequence
+This makes the third promoter test sequence

← Older revision		Revision as of 15:01, 3 July 2008
Line 2:		Line 2:

	The pMnt/TetR promoter is constitutively on and is repressed by the TetR protein. The addition of aTc should repress the TetR causing it to glow. The addition of the Mnt protein should also repress the promoter, causing it to not glow.		The pMnt/TetR promoter is constitutively on and is repressed by the TetR protein. The addition of aTc should repress the TetR causing it to glow. The addition of the Mnt protein should also repress the promoter, causing it to not glow.
		+
		+	{\| border="1" cellpadding="2"
		+	!width="30"\|TetR
		+	!width="30"\|Mnt
		+	!width="30"\|aTc + GFP
		+	!width="30"\|aTc - GFP
		+	\|-
		+	\| 0 \|\| 0 \|\| 1 \|\| 1
		+	\|-
		+	\| 0 \|\| 1 \|\| 0 \|\| 0
		+	\|-
		+	\| 1 \|\| 0 \|\| 1 \|\| 0
		+	\|-
		+	\| 1 \|\| 1 \|\| 0 \|\| 0
		+	\|}
		+

@@ Line 12: / Line 12: @@
 add aTc
-pBAD→RBS+TetR+TT+pMnt/Tet→GFP glows
+pBAD→RBS+TetR+TT+pMnt/Tet→RBS+GFP glows
 I13453+[http://partsregistry.org/Part:BBa_B0034 B0034]+[http://partsregistry.org/Part:BBa_C0040 C0040]+B0015+K091105+E0240
@@ Line 19: / Line 19: @@
 '''2) Test Mnt repression:'''
-pBAD→RBS+Mnt+TT+pMnt/Tet→GFP glows
+pBAD→RBS+Mnt+TT+pMnt/Tet→RBS+GFP glows
-I13453+B0034+[http://partsregistry.org/Part:BBa_C0072 C0072]+B0015+K091105+E0240
+I13453+[http://partsregistry.org/Part:BBa_I13025 I13025]+B0015+K091105+E0240
 '''3) Test Tet and Mnt repression:'''
-pBAD→RBS+TetR+RBS+Mnt+TT+pMnt/Tet→GFP does not glows
+pBAD→RBS+TetR+RBS+Mnt+TT+pMnt/Tet→RBS+GFP does not glows
 I13453+B0034+C0040+B0034+C0072+B0015+K091105+E0240
@@ Line 37: / Line 37: @@
 K091105 ligated with E0240
-need to ligate some promoter (digest with E and S) with an RBS (digest E and X) and ligate TetR(digest with E and S) with a double terminator (digest with E and x)
+need to ligate pBAD to and RBS
-then ligate the promoter+RBS to the 5' end of the TetR+TT parts together
+then ligate TetR to the 3' end of that
 then ligate that part 5' to the pMnt/Tet+GFP
@@ Line 47: / Line 49: @@
-ligate Mnt to the double terminator
+ligate the part RBS+Mnt to the 5' end of the TT
-the ligate the promoter+RBS from the first test sequence to the 5' end of the Mnt+TT
+then ligate pBAD to the 5' end of that
-ligate this to the 5' end of the pMnt/Tet+GFP
+then ligate the part pMnt/Tet+RBS+GFP to the 3' end of that
 This makes the second promoter test sequence.
@@ Line 57: / Line 59: @@
+then ligate the part pMnt/Tet+RBS+GFP to the 3' end of that
 This makes the third promoter test sequence

← Older revision		Revision as of 15:18, 2 July 2008
Line 71:		Line 71:

	the first test sequence should glow and the third should not.		the first test sequence should glow and the third should not.
−
−
−	~~'''Parts Relevant to XOR Gate:'''~~
−
−	~~{\| border="1" cellpadding="2"~~
−	~~!width="30"\|part no.~~
−	~~!width="30"\|description~~
−	~~!width="30"\|source~~
−	~~!width="30"\|responsible~~
−	\|-
−	~~\| K091105 \|\| pMnt/Tet \|\| MW '08 \|\| RC~~
−	\|-
−	~~\| B0034 \|\| RBS \|\| registry \|\| ?~~
−	\|-
−	~~\| C0061 \|\| Lux gene(+LVA) \|\| registry \|\| ?~~
−	\|-
−	~~\| C0062 \|\| Lux gene(-LVA) \|\| registry \|\| ?~~
−	\|-
−	~~\| C0051 \|\| cI gene \|\| registry \|\| ?~~
−	\|-
−	~~\| B0015 \|\| TT \|\| registry \|\| ?~~
−	\|}
−
−
−
−
−
−	~~'''Steps Needed for Test XOR Gate:'''~~
−
−	~~{\| border="1" cellpadding="2"~~
−	~~!width="30"\|step no.~~
−	~~!width="30"\|vector BBa~~
−	~~!width="30"\|vector description~~
−	~~!width="30"\|vector suffix prefix~~
−	~~!width="30"\|vector enzymes~~
−	~~!width="30"\|vector size (bp)~~
−	~~!width="30"\|insert BBa~~
−	~~!width="30"\|insert description~~
−	~~!width="30"\|insert suffix prefix~~
−	~~!width="30"\|insert enzymes~~
−	~~!width="30"\|insert size (bp)~~
−	~~!width="30"\|product BBa~~
−	~~!width="30"\| product size~~
−	\|-
−	~~\| 1 \|\| B0034 \|\| RBS \|\| suffix \|\| SP \|\| 2169 \|\| C0061 \|\| Lux (+LVA) \|\| suffix, prefix \|\| XP \|\| 756 \|\| ? \|\| 3011~~
−	\|-
−	~~\| 2 \|\| B0034 \|\| RBS \|\| suffix \|\| SP \|\| 2169 \|\| C0051 \|\| cI \|\| suffix, prefix \|\| XP \|\| 750 \|\| ? \|\| 3005~~
−	\|-
−	~~\| 3 \|\| product from step 1 \|\| RBS+ Lux \|\| suffix \|\| SP \|\| 3011 \|\| product from step 2 \|\| RBS+ cI \|\| suffix, prefix \|\| XP \|\| 762 \|\| ? \|\| 3773~~
−	\|-
−	~~\| 4 \|\| B0015 \|\| TT \|\| prefix \|\| RX \|\| 2278 \|\| product from step 3 \|\| RBS+Lux+ RBS+cI \|\| suffix, prefix \|\| RS \|\| 3612 \|\| ? \|\| 3894~~
−	\|-
−	~~\| 5 \|\| K091105 \|\| pMnt/Tet \|\| suffix \|\| SP \|\| 2255 \|\| product from step 4 \|\| RBS+Lux+RBS+cI+TT \|\| suffix prefix \|\| XP \|\| 3751 \|\| ? \|\| 3992~~
−	\|-

@@ Line 115: / Line 115: @@
 !width="30"| product size
 |-
-| 1 || B0034 || RBS || suffix || SP || 2100 || C0061 || Lux (+LVA) || suffix, prefix || XP || 756 || ? || 2850
+| 1 || B0034 || RBS || suffix || SP || 2169 || C0061 || Lux (+LVA) || suffix, prefix || XP || 756 || ? || 3011
 |-
-| 2 || B0034 || RBS || suffix || SP || 2100 || C0051 || cI || suffix, prefix || XP || 750 || ? || 2850
+| 2 || B0034 || RBS || suffix || SP || 2169 || C0051 || cI || suffix, prefix || XP || 750 || ? || 3005
 |-
-| 3 || product from step 1 || RBS+ Lux || suffix || SP || 2850 || product from step 2 || RBS+ cI || suffix, prefix || XP || 762 || ? || 3612
+| 3 || product from step 1 || RBS+ Lux || suffix || SP || 3011 || product from step 2 || RBS+ cI || suffix, prefix || XP || 762 || ? || 3773
 |-
-| 4 || product from step 3 || RBS+Lux+ RBS+cI || suffix, prefix || RS || 3612 || B0015 || TT || prefix || RX || 2221 || ? || 3751
+| 4 || B0015 || TT || prefix || RX || 2278 || product from step 3 || RBS+Lux+ RBS+cI || suffix, prefix || RS || 3612 || ? || 3894
 |-
-| 5 || product from step 4 || RBS+Lux+RBS+cI+TT || suffix prefix || XP || 3751 || K091105 || pMnt/Tet || suffix || SP || 2200 || ? || 3949
+| 5 || K091105 || pMnt/Tet || suffix || SP || 2255 || product from step 4 || RBS+Lux+RBS+cI+TT || suffix prefix || XP || 3751 ||   ? || 3992
 |-

@@ Line 73: / Line 73: @@
-'''Parts Needed for XOR Gate:'''
+'''Parts Relevant to XOR Gate:'''
 {| border="1" cellpadding="2"
@@ Line 92: / Line 92: @@
 |-
 | B0015 || TT || registry || ?
 |}