IGEM:Cambridge/2008/Turing Pattern Formation/Vectors: Difference between revisions
From OpenWetWare
Jump to navigationJump to search
(5 intermediate revisions by 2 users not shown) | |||
Line 5: | Line 5: | ||
<hr width="810px"> | <hr width="810px"> | ||
All these vectors were kindly provided by Dr. Dan Ziegler, Director of the Bacillus Genetic Stock Center at Ohio State University | All these vectors were kindly provided by Dr. Dan Ziegler, Director of the Bacillus Genetic Stock Center at Ohio State University. | ||
= Integration Vectors = | = Integration Vectors = | ||
Line 73: | Line 71: | ||
* 4 pstI sites, EcoRI site, no Xba site | * 4 pstI sites, EcoRI site, no Xba site | ||
| <nowiki>*</nowiki>: can use PyrD & AmyE instead | | <nowiki>*</nowiki>: can use PyrD & AmyE instead | ||
| bgcolor="red" fontcolor="white"| No: | | bgcolor="red" fontcolor="white"| No: wrong size | ||
|- | |- | ||
| ECE176 | | ECE176 | ||
Line 90: | Line 88: | ||
* 2 pstI sites, X and S before promoter, E after terminator | * 2 pstI sites, X and S before promoter, E after terminator | ||
| <nowiki>*</nowiki>: use of strong rpsD promoter, but hard to insert into | | <nowiki>*</nowiki>: use of strong rpsD promoter, but hard to insert into | ||
| bgcolor=" | | bgcolor="green" fontcolor="white"| Yes | ||
|- | |- | ||
| ECE153 | | ECE153 | ||
Line 107: | Line 105: | ||
* multiple Xbas, multiple pstIs | * multiple Xbas, multiple pstIs | ||
| <nowiki>***</nowiki>: inducible promoter, strong gfp, no sequence | | <nowiki>***</nowiki>: inducible promoter, strong gfp, no sequence | ||
| bgcolor=" | | bgcolor="green" fontcolor="white"| yes | ||
|- | |- | ||
| ECE162 | | ECE162 | ||
Line 124: | Line 122: | ||
* multiple Xbas, multiple pstIs | * multiple Xbas, multiple pstIs | ||
| <nowiki>***</nowiki>: inducible promoter, strong gfp, no sequence | | <nowiki>***</nowiki>: inducible promoter, strong gfp, no sequence | ||
| bgcolor="yellow" fontcolor="white"| inconclusive | | bgcolor="yellow" fontcolor="white"| inconclusive : only one band | ||
|- | |- | ||
Line 151: | Line 149: | ||
| [[image:ECE149.jpg|thumb|ECE149 Map]] | | [[image:ECE149.jpg|thumb|ECE149 Map]] | ||
| [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pmutingfp.htm From Bayreuth] | | [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pmutingfp.htm From Bayreuth] | ||
| bgcolor="red" fontcolor="white"| No: | | bgcolor="red" fontcolor="white"| No: 3 bands on the gel | ||
|- | |- | ||
| ECE150 | | ECE150 | ||
Line 163: | Line 161: | ||
| [[image:ECE153.jpg|thumb|ECE150 Map]] | | [[image:ECE153.jpg|thumb|ECE150 Map]] | ||
| [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pmutincfp.htm From Bayreuth] <br/> (sequence incomplete) | | [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pmutincfp.htm From Bayreuth] <br/> (sequence incomplete) | ||
| bgcolor=" | | bgcolor="green" fontcolor="white"| yes | ||
|- | |- | ||
| ECE151 | | ECE151 | ||
Line 186: | Line 184: | ||
| [[image:ECE147.jpg|thumb|ECE147 Map]] | | [[image:ECE147.jpg|thumb|ECE147 Map]] | ||
| [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pmutincmyc.htm From Bayreuth] | | [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pmutincmyc.htm From Bayreuth] | ||
| bgcolor=" | | bgcolor="green" fontcolor="white"| yes | ||
|} | |} | ||
Line 206: | Line 204: | ||
* Ampicillin selectable only in EC | * Ampicillin selectable only in EC | ||
* gfpmut3a in front of MCS, no promoter, but gram-positive RBS | * gfpmut3a in front of MCS, no promoter, but gram-positive RBS | ||
| [[image: | | [[image:ECE165.jpg|thumb|ECE165 Map]] | ||
| [http://www.bgsc.org/sequences/pad123.htm BGSC sequence] | | [http://www.bgsc.org/sequences/pad123.htm BGSC sequence] | ||
| bgcolor="green" fontcolor="white"| Yes | | bgcolor="green" fontcolor="white"| Yes | ||
Line 236: | Line 234: | ||
* Ampicillin selectable only in EC | * Ampicillin selectable only in EC | ||
* strong xylose inducible PxylA promoter in front of MCS | * strong xylose inducible PxylA promoter in front of MCS | ||
| [[image: | | [[image:ECE188.jpg|thumb|ECE189 Map]] | ||
| [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pHCMC04.htm From Bayreuth] | | [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pHCMC04.htm From Bayreuth] | ||
| bgcolor="white" fontcolor="white"| Not yet checked, DNA stocks | | bgcolor="white" fontcolor="white"| Not yet checked, DNA stocks | ||
Line 246: | Line 244: | ||
* Ampicillin selectable only in EC | * Ampicillin selectable only in EC | ||
* strong xylose inducible PxylA promoter in front of MCS | * strong xylose inducible PxylA promoter in front of MCS | ||
| [[image: | | [[image:ECE188.jpg|thumb|ECE190 Map]] | ||
| [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pHCMC05.htm From Bayreuth] | | [http://www.genetik.uni-bayreuth.de/LSGenetik1/schumann_pHCMC05.htm From Bayreuth] | ||
| bgcolor=" | | bgcolor="green" fontcolor="white"| Yes | ||
|} | |} | ||
Latest revision as of 04:57, 29 August 2008
All these vectors were kindly provided by Dr. Dan Ziegler, Director of the Bacillus Genetic Stock Center at Ohio State University.
Integration Vectors
These vectors integrate into the chromosome and do not have a replication origin in Bacillus. They either integrate cassettes that require double crossovers and two homologous regions, or whole-plasmid insertions that only require one crossover and one corresponding region of homology.
Ectopic Integration
The integration site of these is pre-determined
BGSC Accession | Vector Name | Features | Integration Locus | Vector Map | Vector Sequence | Possible Uses | Necessity (1-5 stars) | Checked on Gel |
---|---|---|---|---|---|---|---|---|
ECE112 | pDG1661 |
|
amyE | U46196 |
|
**** | Yes | |
ECE171 | pPyr-Kan |
|
pyrD | AY464559 |
|
**** | Yes | |
ECE172 | pGlt-Cm |
|
gltA | AY464560 |
|
*: can use PyrD & AmyE instead | No: wrong size | |
ECE176 | pBCJ164 |
|
rspD | BGSC sequence |
|
*: use of strong rpsD promoter, but hard to insert into | Yes | |
ECE153 | pSG1154 |
|
amyE | not available |
|
***: inducible promoter, strong gfp, no sequence | yes | |
ECE162 | pSG1193 |
|
amyE | not available |
|
***: inducible promoter, strong gfp, no sequence | inconclusive : only one band |
Insert Integration
The user inserts a homologous piece of the Bacillus chromosome and the vector integrates there.
BGSC Accession | Vector Name | Features | Vector Map | Vector Sequence | Checked on gel |
---|---|---|---|---|---|
ECE149 | pMUTIN-GFP+ |
|
From Bayreuth | No: 3 bands on the gel | |
ECE150 | pMUTIN-CFP |
|
From Bayreuth (sequence incomplete) |
yes | |
ECE151 | pMUTIN-YFP |
|
From Bayreuth | Yes | |
ECE147 | pMUTIN-cMyc |
|
From Bayreuth | yes |
Shuttle Vectors
BGSC Accession | Vector Name | Features | Vector Map | Vector Sequence | Checked on gel |
---|---|---|---|---|---|
ECE165 | pAD123 |
|
BGSC sequence | Yes | |
ECE166 | pAD43-25 |
|
BGSC sequence | Yes | |
ECE188 | pHCMC02 |
|
From Bayreuth | Not yet checked, DNA stocks | |
ECE189 | pHCMC04 |
|
From Bayreuth | Not yet checked, DNA stocks | |
ECE190 | pHCMC05 |
|
From Bayreuth | Yes |
Plasmid Gels
- We ran these plasmids on gels, using double and single digests, to check them