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2017-08-28T11:31:06Z

Bzhu:

List of plasmids

2017-08-28T11:27:16Z

Bzhu:

Here you can find a list of ''Bacillus subtilis'' plasmids:

==Fusion to reporter genes==
*[[pAC5]]: translational ''lacZ''-fusions (chloramphenicol resistence)
*[[pAC6]]: transcriptional ''lacZ''-fusions (cat resistence)
*[[pAC7]]: translational ''lacZ''-fusions (kanamycin resistence)
* pBaSysBioII: gfp transcriptional fusions for high-throughput analysis of gene expression {{PubMed|20150235}}
* a suite of vectors for ectopic insertion of GFP, CFP and IYFP transcriptional fusions in single copy at the ''amyE'' and ''bglS'' loci {{PubMed|20600285}}
* List of GFP variants useful for various fusions (IPTG inducible P''hyperspank'' with ''amyE'' integration) {{PubMed|23956387}}

==Labelling of a protein with a flourescent tag==
* [[pBP43]]: integrative plasmid for the fusion of monomeric GFP tag to the C-terminus of a protein, keeping the control of the expression under its natural promoter, '''this is the preferred plasmid for constructing GFP fusions!'''
* [[pGP1871]]: integrative plasmid for the fusion of YFP tag to the C-terminus of a protein, keeping the control of the expression under its natural promoter
* [[pGP1080]]: integrative plasmid for the fusion of GFP tag to the C-terminus of a protein, keeping the control of the expression under its natural promoter, allows express of the downstream gene by the P(spac) promoter, '''ATTENTION: This plasmid encodes the dimeric GFP that tends to produce artifacts!!!'''
* [[pGP1870]]: integrative plasmid for the fusion of GFP tag to the C-terminus of a protein, keeping the control of the expression under its natural promoter, '''ATTENTION: This plasmid encodes the dimeric GFP that tends to produce artifacts!!!'''
* ectopic integration vectors for generating fluorescent promoter fusions in ''Bacillus subtilis'' with minimal dark noise {{PubMed|24874808}}

==Labelling of a protein with a triple FLAG tag==
* [[pGP1087]]: integrative plasmid for the fusion of 3x FLAG tag to the C-terminus of a protein, keeping expression under the control of the natural promoter, allowing the expression of downstream genes under the control of the IPTG-inducible Pspac promoter
* [[pGP1331]]: integrative plasmid for the fusion of 3x FLAG tag to the C-terminus of a protein, keeping expression under the control of the natural promoter
* [[pGP1370]]: replicative plasmid for the fusion of 3x FLAG tag to the C-terminus of a protein, high expression due to the strong ''[[degQ]]''(Hy) promoter from [[pBQ200]]

==Expression/ purification in ''B. subtilis''==
*[[pBP26]]: expression of YFP in ''B. subtilis'' and in ''E. coli''
*[[pBP27]]: expression of CFP in ''B. subtilis'' and in ''E. coli''
*[[pBQ200]]: overexpression of proteins in ''B. subtilis'' under control of a strong ''degQ36'' promoter
*[[pGP380]]: expression of proteins in ''B. subtilis'' allows fusion to a Strep-tag at the N-terminus of the protein
*[[pGP382]]: expression of proteins in ''B. subtilis'' allows fusion to a Strep-tag at the C-terminus of the protein
*[[pGP886]]: integration vector (integrates in ''[[xkdE]]''); allowing the expression of genes under the control of the xylose-inducible PxylA promoter in ''B. subtilis''
*[[pGP888]]: integration vector (integrates in ''[[ganA]]''); allowing the expression of genes under the control of the xylose-inducible PxylA promoter in ''B. subtilis''
*[[pGP1459]]: integration vector (integrates in ''[[ganA]]'') analogous to [[pGP380]]; expression of proteins in ''B. subtilis'' allows fusion to a Strep-tag at the N-terminus of the protein
*[[pGP1460]]: integration vector (integrates in ''[[ganA]]'') analogous to [[pGP382]]; expression of proteins in ''B. subtilis'' allows fusion to a Strep-tag at the C-terminus of the protein
* [[pGP1389]]: integrative plasmid for the fusion of a Strep-tag to the C-terminus of a protein, keeping expression under the control of the natural promoter

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P''grac'' vectors
* [[pHT01]]: Expression vector for high level protein production with ''B. subtilis''. Expression of the gene of interest will be under control of the IPTG inducible Pgrac promoter. ''E.coli / B. subtilis'' shuttle vector (amp/cm)
* [[pHT43]]: Secretion vector, for high level protein production with ''B. subtilis''. The gene of interest will be fused to the signal sequence of ''amyQ'' included in the vector. Expression will be under control of the IPTG inducible P''grac'' promoter . ''E. coli / B. subtilis'' shuttle vector (amp/cm)
* [[pHT08]]: [[pHT01]] variant with N-terminal 8xHis tag fusion
* [[pHT09]]: [[pHT01]] variant with N-terminal Strep tag fusion
* [[pHT10]]: [[pHT01]] variant with C-terminal c-Myc tag fusion

P''grac100'' vectors
* [[pHT253]]:Expression vector for high level protein production with ''B. subtilis''. Expression of the gene of interest will be under control of the IPTG inducible enhanced P''grac'' promoter "P''grac100''". N-terminal His tag; ''E. coli / B. subtilis'' shuttle vector (amp/cm)

* [[pHT254]]: Expression vector for high level protein production with ''B. subtilis''. Expression of the gene of interest will be under control of the IPTG inducible enhanced P''grac'' promoter "P''grac100''". C-terminal His tag; ''E. coli / B. subtilis'' shuttle vector (amp/cm)

* [[pHT255]]:Expression vector for high level protein production with B. subtilis. Expression of the gene of interest will be under control of the IPTG inducible enhanced Pgrac promoter "Pgrac100". C-terminal Strep tag; E. coli / B. subtilis shuttle vector (amp/cm)

Food Grade expression vectors
* [[pTTB1]]: Low copy food grade expression vector; constitutive expression; selection is based on the interplay of the endogenous ''Bacillus'' toxin EndoA (encoded within the genome of host strains ''B. subtilis'' TEA or ''B. subtilis'' WEA) and its antitoxin EndoB (vector encoded); ''B. subtilis / E.coli'' shuttle vector
* [[pTTB2]]: High copy food grade expression vector; constitutive expression; selection is based on the interplay of the endogenous ''Bacillus'' toxin EndoA (encoded within the genome of host strains ''B. subtilis'' TEA or ''B. subtilis'' WEA) and its antitoxin EndoB (vector encoded); ''B. subtilis / E.coli'' shuttle vector
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==Expression/ purification in ''E. coli''==
*[[pWH844]]: expression of proteins in ''E. coli'', allows fusion to a N-terminal His-tag
*[[pGP570]]: expression of proteins in ''E. coli'', allows fusion to a N-terminal His-tag that can be cleaved off by thrombin
*[[pETM-11]]: expression of proteins in ''E. coli'', allows fusion to a N-terminal His-tag that can be cleaved off by TEV protease
*[[pGP172]]: expression of proteins in ''E. coli'' allows fusion to a Strep-tag at the N-terminus of the protein
*[[pGP574]]: expression of proteins in ''E. coli'' allows fusion to a Strep-tag at the C-terminus of the protein

== Construction of (markerless) deletions ==
*[[pMAD]]: construction of markerless KOs in Gram-positive bacteria

== Toolboxes for the work with ''B. subtilis'' ==
* the biobrick box {{PubMed|24295448}}
* a part toolbox to tune genetic expression {{PubMed|27402159}}

PTTB1

2017-08-28T09:26:53Z

Bzhu: