IGEM:Cambridge/2008/Turing Pattern Formation/Vectors

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All these vectors were kindly provided by Dr. Dan Ziegler, Director of the Bacillus Genetic Stock Center at Ohio State University.

Daniel Goodman 17:01, 31 July 2008 (UTC):I just put this table together, but I have yet to add the plasmid maps to these. For now, to see the maps, go to BGSC homepage and find them in the catalog. They are also in the black Turing Group binder under "ordered vectors and strains".

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
ECE112	pDG1661	Chloramphenicol selectable in EC and BS Spectinomycin selectable only in EC spoVG-lacZ fusion: contains spoVG RBS insert must contain a promoter cassette insertion	amyE	map	U46196
ECE171	pPyr-Kan	Kanamycin selectable in EC or BS Ampicillin selectable only in EC cassette insertion	pyrD	map	AY464559
ECE172	pGlt-Cm	Cloramphenicol selectable in EC or BS Ampicillin selectable only in EC cassette insertion	gltA	map	AY464560
ECE176	pBCJ164	Cloramphenicol selectable in EC or BS Ampicillin selectable only in EC cassette insertion contains strong rspD promoter and RBS	rspD	map	BGSC sequence
ECE153	pSG1154	Spectinomycin selectable in EC or BS Ampicillin selectable only in EC cassette insertion xylose-inducible promoter high fluorescence GFP variant for protein fusions	amyE	map	not available
ECE162	pSG1193	Spectinomycin selectable in EC or BS Ampicillin selectable only in EC YFP full coding sequence, including RBS and Pxyl xylose-inducible cassette insertion	amyE	map	not available

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
ECE149	pMUTIN-GFP+	Erythromycin selectable in EC or BS Ampicillin selectable only in EC GFP+ full coding sequence, including RBS and Pspac IPTG-inducible full plasmid insertion	map	From Bayreuth
ECE150	pMUTIN-CFP	Erythromycin selectable in EC or BS Ampicillin selectable only in EC CFP full coding sequence, including RBS and Pspac IPTG-inducible full plasmid insertion	map	From Bayreuth (sequence incomplete)
ECE151	pMUTIN-YFP	Erythromycin selectable in EC or BS Ampicillin selectable only in EC YFP full coding sequence, including RBS and Pspac IPTG-inducible full plasmid insertion	map	From Bayreuth
ECE147	pMUTIN-cMyc	Erythromycin selectable in EC or BS Ampicillin selectable only in EC cMyc antibody tag fusion IPTG-inducible Pspac promoter	map	From Bayreuth

Shuttle Vectors

BGSC Accession	Vector Name	Features	Vector Map	Vector Sequence
ECE165	pAD123	Chloramphenicol selectable in EC or BS Ampicillin selectable only in EC gfpmut3a in front of MCS, no promoter, but gram-positive RBS	map	BGSC sequence
ECE166	pAD43-25	Chloramphenicol selectable in EC or BS Ampicillin selectable only in EC gfpmut3a in front of MCS, strong Pupp promoter, gram-positive RBS	map	BGSC sequence
ECE188	pHCMC02	Chloramphenicol selectable in EC or BS Ampicillin selectable only in EC weak constitutive PlepA promoter in front of MCS	map	From Bayreuth
ECE189	pHCMC04	Chloramphenicol selectable in EC or BS Ampicillin selectable only in EC strong xylose inducible PxylA promoter in front of MCS	map	From Bayreuth
ECE190	pHCMC05	Chloramphenicol selectable in EC or BS Ampicillin selectable only in EC strong xylose inducible PxylA promoter in front of MCS	map	From Bayreuth

Plasmid Gels

• We ran these plasmids on gels, using double and single digests, to check them