IGEM:IMPERIAL/2007/DNA Constructs: Difference between revisions

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|width="--"%|Application
|width="--"%|Application
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||  ||  || [http://parts.mit.edu/registry/index.php/Part:BBa_I719021 BBa_I719021] || ptet & luxI || ''Paused'' || [http://parts.mit.edu/registry/index.php/Part:BBa_R0040 R0040] + [http://parts.mit.edu/registry/index.php/Part:BBa_F1610 F1610] || ID final sender (''duplicated'' - [http://parts.mit.edu/registry/index.php/Part:BBa_J23039 J23039])
||  ||  || [http://parts.mit.edu/registry/index.php/Part:BBa_I719021 BBa_I719021] || ptet & luxI || ''Deleted'' || [http://parts.mit.edu/registry/index.php/Part:BBa_R0040 R0040] + [http://parts.mit.edu/registry/index.php/Part:BBa_F1610 F1610] || ID final sender (''duplicated'' - [http://parts.mit.edu/registry/index.php/Part:BBa_J23039 J23039])


|- style="background:#CCFFCC" align="center"
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||  ||  || [http://parts.mit.edu/registry/index.php/Part:BBa_I719022 BBa_I719022] || pcI & GFP || Planning || [http://parts.mit.edu/registry/index.php/Part:BBa_R0051 BBa_R0051] +  [http://parts.mit.edu/registry/index.php/Part:BBa_I13504 BBa_I13504] || CBD test construct (pcI)
||  ||  || [http://parts.mit.edu/registry/index.php/Part:BBa_I719022 BBa_I719022] || pcI & GFP || ''Paused''|| [http://parts.mit.edu/registry/index.php/Part:BBa_R0051 BBa_R0051] +  [http://parts.mit.edu/registry/index.php/Part:BBa_I13504 BBa_I13504] || CBD test construct (pcI)


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|- style="background:#CCFFCC" align="center"
|- style="background:#CCFFCC" align="center"
||  ||  || BBa_I719??? || pT7 & DsRed || Pending || pT7 + DsRed || CBD final construct (pT7)
||  ||  || BBa_I719??? || pT7 & DsRed || Planning || pT7 + DsRed || CBD final construct (pT7)


|- style="background:#CCFFCC" align="center"
|- style="background:#CCFFCC" align="center"
||  ||  || BBa_I719??? || ptet & DsRed || Pending || [http://parts.mit.edu/registry/index.php/Part:BBa_R0040 BBa_R0040]+ DsRed || CBD final construct (ptet)
||  ||  || BBa_I719??? || ptet & DsRed || Planning || [http://parts.mit.edu/registry/index.php/Part:BBa_R0040 BBa_R0040] + DsRed || CBD final construct (ptet)


|- style="background:#CCFFCC" align="center"
|- style="background:#CCFFCC" align="center"
||  ||  || BBa_I719??? || pcI-DsRed || Pending || [http://parts.mit.edu/registry/index.php/Part:BBa_R0051 BBa_R0051] + DsRed || CBD final construct (pcI)
||  ||  || BBa_I719??? || pcI-DsRed || ''Paused'' || [http://parts.mit.edu/registry/index.php/Part:BBa_R0051 BBa_R0051] + DsRed || CBD final construct (pcI)


|- style="background:#CCFFCC" align="center"
|- style="background:#CCFFCC" align="center"
||  ||  || BBa_I719??? || ptet-luxR-plux-DsRed || Pending || [http://parts.mit.edu/registry/index.php/Part:BBa_F2620 BBa_F2620] + DsRed || ID detector final construct
||  ||  || BBa_I719??? || ptet-luxR-plux-DsRed || Pending || [http://parts.mit.edu/registry/index.php/Part:BBa_T2620 BBa_F2620] + DsRed || ID detector final construct 1


|- style="background:#CCFFCC" align="center"
|- style="background:#CCFFCC" align="center"
||  ||  ||  ||  ||  ||  ||
||  ||  || BBa_I719??? || plux-DsRed || Pending || [http://parts.mit.edu/registry/index.php/Part:BBa_R0062 BBa_R0062] + DsRed || ID detector final construct 2
 
|- style="background:#CCFFCC" align="center"
||  ||  || BBa_I719??? || hrpR-hrpS & phrpL|| Pending ||hrpR-hrpS + phrpL || ID detector test construct 3 and final construct 5
 
|- style="background:#CCFFCC" align="center"
||  ||  || BBa_I719??? || Ptet-luxR-Plux & hrpR-hrpS-phrpL || Pending || [http://parts.mit.edu/registry/index.php/Part:BBa_F2620 BBa_F2620] + hrpR-hrpS-phrpL || ID detector test construct 3 and final construct 5
 
|- style="background:#CCFFCC" align="center"
||  ||  || BBa_I719??? || Ptet-luxR-Plux-hrpR-hrpS-phrpL & GFP || Pending ||Ptet-luxR-Plux-hrpR-hrpS-phrpL + [http://parts.mit.edu/registry/index.php/Part:BBa_E0240 BBa_E0240] || ID detector test construct 3
 
|- style="background:#CCFFCC" align="center"
||  ||  || BBa_I719??? || Ptet-luxR-Plux-hrpR-hrpS-phrpL & DsRed|| Pending ||Ptet-luxR-Plux-hrpR-hrpS-phrpL + DsRed || ID detector final construct 5


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==Construct Diagrams==
==Construct Diagrams==
The original (old) diagrams page was moved to [[IGEM:IMPERIAL/2007/DNA_Constructs/Con_Diagrams_old|old construct diagrams]] page.
The original (old) diagrams page was moved to [[IGEM:IMPERIAL/2007/DNA_Constructs/Con_Diagrams_old|old construct diagrams]] page.  This page also contains the obsolete parts.


==Constructs for Cell By Date==
===CBD Constructs===
'''ptet GFP'''
[[image:ICGEMS_dvc_Ptet_gfp.png|left|thumb|200px|Ptet promoting GFP]]
*Availability: Found in registry as [http://parts.mit.edu/registry/index.php/Part:BBa_I13522 BBa_I13522]
<br clear="all">


*Construct CBD 1
'''pT7 GFP'''
**Entire construct is currently available in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_E7104 BBa_E7104] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
[[image:ICGEMS_dvc_PT7_gfp.png|left|thumb|200px|PT7 promoting GFP]]
**pT7 can be used with T7 RNAP in vitro and in veso, and it gives constitutive expression.
*Availability: Found in resigry as [http://parts.mit.edu/registry/index.php/Part:BBa_E7104 BBa_E7104]  
[[image:ICGEMS_dvc_PT7_gfp.png|left|thumb|200px|PT7 promoting GFP]]<br clear="all">
<br clear="all">
 
*Construct CBD 2
**Entire construct is currently in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_I13522 BBa_I13522] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
**pTet can be used with E.coli RNAP in vitro and maybe in veso, and it gives constitutive expression.
[[image:ICGEMS_dvc_Ptet_gfp.png|left|thumb|200px|Ptet promoting GFP]]<br clear="all">
 
*Construct CBD 3
**pBAD promoter with GFP is currently available in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_J5528 BBa_J5528] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB2K3 pSB2K3] (KanR)
**Need Ara C regulator protein in vitro and in veso. Ara C available with the RBS:[http://parts.mit.edu/registry/index.php/Part:BBa_S03550 BBa_S03550]
**pBad can be used with E.coli RNAP in vitro, and it is inducible by arabinose.
**To be used in veso, arabinose transport proteins have to be inserted in the phospholipid bilayer of the vesicles.
[[image:ICGEMS_dvc_PBad_gfp.png|left|thumb|400px|PBAD promoting GFP and Ptet promoting araC]]<br clear="all">
 
*Construct CBD 4
**Entire construct is currently available in Biobricks Registry:  [http://parts.mit.edu/registry/index.php/Part:BBa_J07037 BBa_J07037] (avaiable), in [http://parts.mit.edu/registry/index.php/Part:pSB2K3 pSB2K3] (KanR)
**pcI gives constitutive expression.
[[image:PcI_construct.PNG|left|thumb|400px|PcI promoting GFP]]<br clear="all">
 
*Construct CBD 5
**All parts are currently available in the Biobricks Registry.
**pcI promoter: [http://parts.mit.edu/registry/index.php/Part:BBa_R0051 BBa_R0051] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
**Arabinose C regulator protein with RBS: [http://parts.mit.edu/registry/index.php/Part:BBa_S03550 BBa_S03550] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1AC3 pSB1AC3] (A+CR)
**Stop codon: [http://parts.mit.edu/registry/index.php/Part:BBa_B0015 BBa_B0015], in [http://parts.mit.edu/registry/index.php/Part:pSB1AK3 pSB1AK3] (A+KR)
**pBAD with GFP: [http://parts.mit.edu/registry/index.php/Part:BBa_J5528 BBa_J5528], in [http://parts.mit.edu/registry/index.php/Part:pSB2K3 pSB2K3] (KanR)
[[image:ICGEMS_dvc_PBad2_gfp.PNG‎ |left|thumb|400px|PBAD promoting GFP and PcI promoting araC]] <br clear="all">


*Construct CBD 6
'''ptet DsRed express'''
**T7 promoter is in the process of cloning
**dsRed has been ordered
[[Image:ICGEMS_dvc_PT7_rfp.PNG|left|thumb|200px|pT7 promoting dsRed]] <br clear="all">
 
*Construct CBD 7
**tet promoter available in the registry: [http://parts.mit.edu/registry/index.php/Part:BBa_R0040 BBa_R0040]
**dsRed has been ordered
[[image:ICGEMS_dvc_Ptet_rfp.PNG|left|thumb|200px|Ptet promoting dsRed]]
[[image:ICGEMS_dvc_Ptet_rfp.PNG|left|thumb|200px|Ptet promoting dsRed]]
 
*Availability: Requires ligation
*Promoter can be found in registry as: [http://parts.mit.edu/registry/index.php/Part:BBa_R0040 BBa_R0040]
*DsRed express is to be ordered from GeneArt
<br clear="all">
<br clear="all">
*Construct CBD 8
**pBad promoter avaiable in the registry: [http://parts.mit.edu/registry/index.php/Part:BBa_I0500 BBa_I0500]
**dsRed has been ordered
**Arabinose C regulator protein with RBS: [http://parts.mit.edu/registry/index.php/Part:BBa_S03550 BBa_S03550] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1AC3 pSB1AC3] (A+CR)
**Stop codon: [http://parts.mit.edu/registry/index.php/Part:BBa_B0015 BBa_B0015], in [http://parts.mit.edu/registry/index.php/Part:pSB1AK3 pSB1AK3] (A+KR)
**tet promoter available in the registry: [http://parts.mit.edu/registry/index.php/Part:BBa_R0040 BBa_R0040]
[[image:ICGEMS_dvc_PBad_rfp.PNG‎|left|thumb|400px|PBAD promoting RFP and Ptet promoting araC]]


'''pT7 DsRed express'''
[[Image:ICGEMS_dvc_PT7_rfp.PNG|left|thumb|200px|pT7 promoting dsRed]]
*Availability: Requires ligation
*Promoter is being PCRed from plasmid derived externally
*DsRed express is to be ordered from GeneArt
<br clear="all">
<br clear="all">
*Construct CBD 9
**&lambda;cI regulated promoter available in the registry:[http://parts.mit.edu/registry/index.php/Part:BBa_R0051 BBa_R0051]
[[image:ICGEMS_dvc_PcI_rfp.PNG|left|thumb|200px|pcI promoting RFP]]


===ID Constructs===
'''pLux GFP with LuxR'''
[[image:ICGEMS_sys_Ptet_luxR_Plux_gfp.png|left|thumb|400px|Ptet promoting LuxR, Plux promoting GFP]]
*Availability: Found in registry as [http://parts.mit.edu/registry/index.php/Part:BBa_T9002 BBa_T9002]
<br clear="all">
<br clear="all">
*Construct CBD 10
**Parts avaiable ....
[[Image:ICGEMS_dvc_PBad2_rfp.PNG‎|left|thumb|400px|PBAD promoting RFP and PcI promoting araC]]


'''pLux GFP with LuxR'''
[[image:ICGEMS_sys_Plux_gfp.png‎|left|thumb|200px|Plux promoting GFP]]
*Availability: Found in registry as [http://parts.mit.edu/registry/index.php/Part:BBa_J37032 BBa_J37032]
<br clear="all">
<br clear="all">


==Constructs for Infector Detector==
''<<Two more constructs with DsRed express yet to be included>>''
 
*Construct ID 1
**Entire part minus promoter is currently in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_F1610 BBa_F1610] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1AK3 pSB1AK3] (AmpR, KanR)
**pTet is currently in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_R0040 BBa_R0040] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
**pTet can be used with E.coli RNAP in vitro and maybe in veso, and it gives constitutive expression.
[[image:ICGEMS_dvc_Ptet_luxI.png|left|thumb|200px|Ptet promoting LuxI]]<br clear="all">
 
*Construct ID 2
**Entire part is currently in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_J24677 BBa_J24677] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
**Entire part minus GFP reporter system is currently in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_F2620 BBa_F2620] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
**GFP reporter system is currently in Biobricks Registry: BBa_I13504 (available)
**pLux can be used with E.coli RNAP in vitro and maybe in veso, and it is repressed by LuxR but activated by AHL:LuxR complex.
[[image:ICGEMS_sys_Ptet_luxR_Plux_gfp.png|left|thumb|400px|Ptet promoting LuxR, Plux promoting GFP]]<br clear="all">
 
*Construct ID 3
**Entire part minus Hrp system is currently in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_F2620 BBa_F2620](available) in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
**pHrpL can be used with E.coli RNAP in vitro and maybe in veso, and it is inducible by HrpR:HrpS:Sigma 54 complex.
[[Image:ICGEMS_sys_Ptet_luxR_Plux_hrpR-hrpS-gfp.PNG‎ |left|thumb|600px|Ptet promoting LuxR, Plux promoting HrpR and HrpS and PHrpL promoting GFP]]<br clear="all">
 
*Construct ID 4
**Entire part minus dsRed reporter system is currently in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_F2620 BBa_F2620] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
**dsRed has been ordered
[[Image:ICGEMS_sys_Ptet_luxR_Plux_lacz.PNG‎|left|thumb|400px|Ptet promoting LuxI]]<br clear="all">
 
*Construct ID 5
**Entire part minus the hrp system and dsRed reporter system is currently in Biobricks Registry: [http://parts.mit.edu/registry/index.php/Part:BBa_F2620 BBa_F2620] (available), in [http://parts.mit.edu/registry/index.php/Part:pSB1A2 pSB1A2] (AmpR)
**dsRed has been ordered
**hrpR-hrpS-pHrpL has been ordered
[[Image:ICGEMS_sys_Ptet_luxR_Plux_hrpR-hrpS-lacZ.PNG‎ ‎|left|thumb|600px|Ptet promoting LuxR, Plux promoting HrpR and HrpS and PHrpL promoting lacZ]]<br clear="all">

Revision as of 12:28, 30 August 2007

Constructs Overview

Here are several tables listing the bricks which we are cloning, constructing and using, with their relevant details.

Cloning Table

Status of biobricks cloned and their applications:

I ID Part Description Status Ligation Target Use in Constructs
02 BBa_I13522 ptet-GFP XL1, Midi, Mini, BL21 --- Basic construct for chassis characterization
11 BBa_E7104 pT7 GFP XL1, Midi, Mini, BL21 --- CBD test construct (pT7)
I 18 BBa_T9002 ptet-luxR-plux-GFP XL1, Midi, BL21 --- Basic construct for chassis characterization
05 BBa_R0040 ptet XL1, Midi, Mini DsRed CBD final (ptet), ID AHL sender
BBa_I719??? pT7 Pending DsRed CBD final construct (pT7)
07 BBa_F2620 ptet-luxR-plux XL1, Midi, Mini DsRed ID final construct
14 BBa_R0051 pcI Problematic DsRed CBD final construct (pcI)
21 BBa_J23039 ptet-luxI XL1, Midi --- ID sender construct
24 BBa_R0062 plux XL1, Midi --- ID construct 2
25 BBa_J37032 plux-GFP XL1, Midi --- ID construct 2
26 BBa_S03119 ptet-luxR XL1, Midi --- ---
13 BBa_B0015 Double terminator XL1, Midi, Mini --- General part construction
17 BBa_I13507 RFP XL1, Midi --- Not being used anymore
19 BBa_B0034 strong RBS XL1, Waiting Midi --- General part construction
23 BBa_I13504 GFP XL1, Midi pcI General part construction
BBa_I719??? DsRed Ordered ptet, plux, pcI Reporter system for ID, Hrp
BBa_I719??? HrpR-HrpS-pHrpL Ordered ??? Hrp regulators
BBa_I719??? pHrpL Ordered ??? Hrp promoter
BBa_I719??? HrpV Ordered ??? Hrp inhibitor system

Ligation Table

Status of parts ligated and their applications:

I ID Part Description Status Parents Application
BBa_I719021 ptet & luxI Deleted R0040 + F1610 ID final sender (duplicated - J23039)
BBa_I719022 pcI & GFP Paused BBa_R0051 + BBa_I13504 CBD test construct (pcI)
BBa_I719??? pT7 & DsRed Planning pT7 + DsRed CBD final construct (pT7)
BBa_I719??? ptet & DsRed Planning BBa_R0040 + DsRed CBD final construct (ptet)
BBa_I719??? pcI-DsRed Paused BBa_R0051 + DsRed CBD final construct (pcI)
BBa_I719??? ptet-luxR-plux-DsRed Pending BBa_F2620 + DsRed ID detector final construct 1
BBa_I719??? plux-DsRed Pending BBa_R0062 + DsRed ID detector final construct 2

Legend: I (Information), [Status: XL1 - cloned in XL1-Blue; Midi - midiprepped; Mini - miniprepped; BL21 - cloned in BL21(DE3)]

Parts from the Registry

Physical information of biobricks:

A W G S ID Name Description Plasmid Resist Copy Size(B) Size(T) Location Conc.
A W G 02 BBa_I13522 GFP under tetracyclin promoter pSB1A2 AmpR High 937bp 3016bp Well 15H, Plate 2 110/100
A W G 05 BBa_R0040 tetracyclin promoter pSB1A2 AmpR High 54bp 2133bp Well 7O, Plate 1 60
A W G 07 BBa_F2620 lux promoter, with luxR pSB1A2 AmpR High 1061bp 3140bp Well 6B, Plate 1 80
A X G 11 BBa_E7104 T7 GFP pSB1A2 AmpR High 826bp 2905bp Well 13F, Plate 2 100
A W G 13 BBa_B0015 Double terminator pSB1AK3 A+KR High 129bp 3316bp Well 1B, Plate 2 100
W X 14 BBa_R0051 λcI promoter pSB1A2 AmpR High 49bp 2128bp Well 9C, Plate 1 --
A G 17 BBa_I13507 RFP pSB1A2 AmpR High 861bp 2940bp Well 16N, Plate 1 80
A W G 18 BBa_T9002 GFP under lux promoter with LuxR pSB1A3 AmpR High 1945bp 4102bp Well 19B, Plate 2 100
A W 19 BBa_B0034 RBS pSB1A2 AmpR High 12bp 2091bp Well 3O, Plate 1 --
A G 21 BBa_J23039 Constitutive luxI pSB1A2 AmpR High 860bp 2939bp Well 11K, Plate 3 140
A W G 23 BBa_I13504 GFP pSB1A2 AmpR High 875bp 2954bp Well 16E, Plate 1 120
A G 24 BBa_R0062 lux promoter pSB1A2 AmpR High 55bp 2134bp Well 9G, Plate 1 90
A G 25 BBa_J37032 GFP under lux promoter pSB1A2 AmpR High 938bp 3017bp Well 8D, Plate 3 100
A G 26 BBa_S03119 constitutive LuxR pSB1A2 AmpR High 998bp 3077bp Well 19J, Plate 1 60

Our Parts

Physical information of our ligated parts:

A W G S ID Name Description Plasmid Resist Copy Size(B) Size(P) Stock
BBa_I719022 cI with GFP pSB1A2 AmpR High 933bp 3988bp
BBa_I791021 luxI coding region under ptet pSB1A2 AmpR High 860bp 2939bp
BBa_I791??? DsRed under ptet pSB1A2 AmpR High

Legend: A (Available), W (Working) [W - working in vivo, W - working in vitro, X - faulty], G (Checked on gel) [G - incorrect size; X - does not appear on gel], S (Sequenced)
Concentrations are given in ng/μl

Part Graveyard

General information on parts we do not like...

A W ID Name Description Plasmid Size Status Notes
A W 01 BBa_E0040 GFP pSB1A2 720bp XL1, Mini Redundant
A 03 BBa_J5528 pBad-GFP pSB2K3 2093bp failed-XL1 ara construct abandoned
A 04 BBa_J24677 plux-GFP pSB1A2 1787bp XL1, Mini Wrong GFP ligated
A 06 BBa_I0500 pBad pSB2K3 1210bp XL1, Mini, Midi ara construct abandoned
A W 08 BBa_F1610 luxI pSB1AK3 798bp XL1, Mini, Midi Substitute part J23039
A W 09 BBa_R0010 lacI-plac pSB1A2 200bp XL1, Mini Lac construct abandoned
A W 10 BBa_E1010 RFP pSB2K3 681bp XL1, Mini Substitute part I13507
A 12 BBa_S03550 araC pSB1AC3 958bp XL1, Mini, Midi araC construct abandoned
A 15 BBa_J07037 cI-GFP pSB2K3 912bp XL1, Mini No RBS present on this part
A W 16 BBa_E0240 GFP pSB1A2 876bp XL1, Midi Redundant
20 --- AraC pSB1AC3 1095bp Planned araC construct abandoned


Construct Diagrams

The original (old) diagrams page was moved to old construct diagrams page. This page also contains the obsolete parts.

CBD Constructs

ptet GFP

Ptet promoting GFP


pT7 GFP

PT7 promoting GFP


ptet DsRed express

Ptet promoting dsRed
  • Availability: Requires ligation
  • Promoter can be found in registry as: BBa_R0040
  • DsRed express is to be ordered from GeneArt


pT7 DsRed express

pT7 promoting dsRed
  • Availability: Requires ligation
  • Promoter is being PCRed from plasmid derived externally
  • DsRed express is to be ordered from GeneArt


ID Constructs

pLux GFP with LuxR

Ptet promoting LuxR, Plux promoting GFP


pLux GFP with LuxR

Plux promoting GFP


<<Two more constructs with DsRed express yet to be included>>

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