Difference between revisions of "List of plasmids"

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* pBaSysBioII: gfp transcriptional fusions for high-throughput analysis of gene expression {{PubMed|20150235}}
 
* 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}}
 
* 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==
 
==Labelling of a protein with a flourescent tag==
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==Labelling of a protein with a triple FLAG tag==
 
==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
 
* [[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
−
* [[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
+
* [[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''==
 
==Expression/ purification in ''B. subtilis''==
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*[[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
 
*[[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
 
* [[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
 +
*[[pGP3273]]: integrative expression vector for ''B. subtilis'' (integrates in ''[[amyE]]''); allowing the expression of genes under the control of the IPTG-inducible Pgrac promoter, ''E.coli / B. subtilis'' shuttle vector (amp/cm)
 +
*[[pGP3232]]: N-term STRP 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)
 +
*[[pGP3299]]: C-term 6xHIS tag 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)
 +
<div style="border: 1px dashed #aaa; margin: 5px -5px; padding: 0 5px">
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<b>P''grac'' vectors</b>
 +
* [[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
 +
 +
<b>P''grac100'' vectors</b>
 +
* [[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)
 +
 +
<b>Food Grade expression vectors</b>
 +
* [[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
 +
<p style="float:right; font-weight:bold; color: navy; padding:5px">Content from [https://www.mobitec.com/cms/index.html MoBiTec GmbH] [[File:Mobiteclogo.png|40px]] </p>
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<p style="clear:both;margin:0;padding:0"></p>
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</div>
  
 
==Expression/ purification in ''E. coli''==
 
==Expression/ purification in ''E. coli''==
 
*[[pWH844]]: expression of proteins in ''E. coli'', allows fusion to a N-terminal His-tag
 
*[[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
+
*[[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
 
*[[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
 
*[[pGP574]]: expression of proteins in ''E. coli'' allows fusion to a Strep-tag at the C-terminus of the protein
 +
*[[pGP3223]]: expression of proteins in ''E. coli'' allows fusion to OsmY-secretion tag at the C-terminus of the protein
  
 
== Construction of (markerless) deletions ==
 
== Construction of (markerless) deletions ==
 
*[[pMAD]]: construction of markerless KOs in Gram-positive bacteria
 
*[[pMAD]]: construction of markerless KOs in Gram-positive bacteria
  
−
== A collection of biobricks for the work with ''B. subtilis'' ==
+
== Toolboxes for the work with ''B. subtilis'' ==
 
* the biobrick box {{PubMed|24295448}}
 
* the biobrick box {{PubMed|24295448}}
 +
* a part toolbox to tune genetic expression {{PubMed|27402159}}

Latest revision as of 09:14, 26 June 2019

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
  • a suite of vectors for ectopic insertion of GFP, CFP and IYFP transcriptional fusions in single copy at the amyE and bglS loci PubMed
  • List of GFP variants useful for various fusions (IPTG inducible Phyperspank with amyE integration) PubMed

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

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
  • pGP3273: integrative expression vector for B. subtilis (integrates in amyE); allowing the expression of genes under the control of the IPTG-inducible Pgrac promoter, E.coli / B. subtilis shuttle vector (amp/cm)
  • pGP3232: N-term STRP 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)
  • pGP3299: C-term 6xHIS tag 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)

Pgrac 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 Pgrac 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

Pgrac100 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 Pgrac promoter "Pgrac100". 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 Pgrac promoter "Pgrac100". 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

Content from MoBiTec GmbH Mobiteclogo.png

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
  • pGP3223: expression of proteins in E. coli allows fusion to OsmY-secretion 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
  • a part toolbox to tune genetic expression PubMed
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