Difference between revisions of "Part:BBa K4274009"
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sfp gene is a biobrick part from <i>B. amyloliquefaciens</i> FZB42 encoding 4'-phosphopantetheinyl transferase which functions as a primer of nonribosomal peptide synthesis via phosphopantetheinylation of thiotemplates. However, compared to the sfp gene in <i>Bacillus subtilis</i> 168, they only had an amino acid homology of 70%. And the mutation existed in <i>Bacillus subtilis</i> 168 resulted in its unable to produce any lipopeptide biosurfactant. Hence, to allow <i>B. subtilis</i> to produce fengycins, it is essential to first knock-out the mutant gene and then knock-in the correct sfp (Part: BBa_K4274009) and degQ (Part: BBa_K4274010).  
 
sfp gene is a biobrick part from <i>B. amyloliquefaciens</i> FZB42 encoding 4'-phosphopantetheinyl transferase which functions as a primer of nonribosomal peptide synthesis via phosphopantetheinylation of thiotemplates. However, compared to the sfp gene in <i>Bacillus subtilis</i> 168, they only had an amino acid homology of 70%. And the mutation existed in <i>Bacillus subtilis</i> 168 resulted in its unable to produce any lipopeptide biosurfactant. Hence, to allow <i>B. subtilis</i> to produce fengycins, it is essential to first knock-out the mutant gene and then knock-in the correct sfp (Part: BBa_K4274009) and degQ (Part: BBa_K4274010).  
  
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===Usage and Biology===
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==Usage and Biology==
 
Our sfp gene (Gene ID: 45022253) is a biobrick part derived from <i>B. amyloliquefaciens</i> FZB42 encoding 4'-phosphopantetheinyl transferase. As mentioned above, the correct sfp gene is the core of  lipopeptide biosurfactant production, thus the mutant sfp gene in <i>B. subtilis</i> is invalid for producing fengycins.
 
Our sfp gene (Gene ID: 45022253) is a biobrick part derived from <i>B. amyloliquefaciens</i> FZB42 encoding 4'-phosphopantetheinyl transferase. As mentioned above, the correct sfp gene is the core of  lipopeptide biosurfactant production, thus the mutant sfp gene in <i>B. subtilis</i> is invalid for producing fengycins.
  
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Edting the genome of <i>Bacillus subtilis</i> 168 to enable it to produce fengycins. (a) Plasmid design for knocking out the invalid sfp gene of <i>Bacillus subtilis</i> 168 and knocking in the sfp gene from B. amyloliquefaciens FZB42 and degQ gene from <i>Bacillus subtilis</i> 168. (b) Electrophoresis results show that gene editing is successful. (c) Protocol about transformation, induction and elimination of pJOE8999 plasmid in <i>Bacillus subtilis</i> 168. ]]
 
Edting the genome of <i>Bacillus subtilis</i> 168 to enable it to produce fengycins. (a) Plasmid design for knocking out the invalid sfp gene of <i>Bacillus subtilis</i> 168 and knocking in the sfp gene from B. amyloliquefaciens FZB42 and degQ gene from <i>Bacillus subtilis</i> 168. (b) Electrophoresis results show that gene editing is successful. (c) Protocol about transformation, induction and elimination of pJOE8999 plasmid in <i>Bacillus subtilis</i> 168. ]]
  
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[[Image:Parts-keystone-fengycin2.tiff|thumbnail|750px|center|'''Figure 1:'''
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==Source==
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Edting the genome of <i>Bacillus subtilis</i> 168 to enable it to produce fengycins. (a) Plasmid design for knocking out the invalid sfp gene of <i>Bacillus subtilis</i> 168 and knocking in the sfp gene from B. amyloliquefaciens FZB42 and degQ gene from <i>Bacillus subtilis</i> 168. (b) Electrophoresis results show that gene editing is successful. (c) Protocol about transformation, induction and elimination of pJOE8999 plasmid in <i>Bacillus subtilis</i> 168. ]]
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===Source===
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<i>B. amyloliquefaciens</i> FZB42  
 
<i>B. amyloliquefaciens</i> FZB42  
  
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===Sequence and Features===
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==Sequence and Features==
 
<partinfo>BBa_K4274009 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K4274009 SequenceAndFeatures</partinfo>
  
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===References===
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==References==
 
[1]Chen X.H., Koumoutsi A., Scholz R., et al. More than Anticipated – Production of Antibiotics and Other Secondary Metabolites by Bacillus amyloliquefaciens FZB42. Mircrobial Biotech. 16 (2), 14-24 (2009). https://doi.org/10.1159/000142891.
 
[1]Chen X.H., Koumoutsi A., Scholz R., et al. More than Anticipated – Production of Antibiotics and Other Secondary Metabolites by Bacillus amyloliquefaciens FZB42. Mircrobial Biotech. 16 (2), 14-24 (2009). https://doi.org/10.1159/000142891.
  

Revision as of 07:53, 12 October 2022

sfp

sfp gene is a biobrick part from B. amyloliquefaciens FZB42 encoding 4'-phosphopantetheinyl transferase which functions as a primer of nonribosomal peptide synthesis via phosphopantetheinylation of thiotemplates. However, compared to the sfp gene in Bacillus subtilis 168, they only had an amino acid homology of 70%. And the mutation existed in Bacillus subtilis 168 resulted in its unable to produce any lipopeptide biosurfactant. Hence, to allow B. subtilis to produce fengycins, it is essential to first knock-out the mutant gene and then knock-in the correct sfp (Part: BBa_K4274009) and degQ (Part: BBa_K4274010).

Usage and Biology

Our sfp gene (Gene ID: 45022253) is a biobrick part derived from B. amyloliquefaciens FZB42 encoding 4'-phosphopantetheinyl transferase. As mentioned above, the correct sfp gene is the core of lipopeptide biosurfactant production, thus the mutant sfp gene in B. subtilis is invalid for producing fengycins.

To construct a strain of B. subtilis to produce fengycins, we will use sfp_target (gRNA) (part: BBa_K4274008) to target the region of mutant sfp in B. subtilis for knock-out, simultaneously sfp (Part: BBa_K4274009) and degQ (Part: BBa_K4274010) gene were knocked-in in situ. It was used in the composite part PvanP*-sfp_target-sfp_HA_US-p43-K4274013-sfp-K4274014-degQ-sfp_HA_DS (Part: BBa_K4274035) to realize fengycins’ production in B. subtilis.

Figure 1: Edting the genome of Bacillus subtilis 168 to enable it to produce fengycins. (a) Plasmid design for knocking out the invalid sfp gene of Bacillus subtilis 168 and knocking in the sfp gene from B. amyloliquefaciens FZB42 and degQ gene from Bacillus subtilis 168. (b) Electrophoresis results show that gene editing is successful. (c) Protocol about transformation, induction and elimination of pJOE8999 plasmid in Bacillus subtilis 168.

Source

B. amyloliquefaciens FZB42

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 38
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 38
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 38
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 38
    Illegal NgoMIV site found at 95
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 661

References

[1]Chen X.H., Koumoutsi A., Scholz R., et al. More than Anticipated – Production of Antibiotics and Other Secondary Metabolites by Bacillus amyloliquefaciens FZB42. Mircrobial Biotech. 16 (2), 14-24 (2009). https://doi.org/10.1159/000142891.

[2]Jin P., Wang H., Liu W., et al. Characteriztion of IpaH2 gene corresponding to lipopeptide synthesis in Bacillus amyloliquefaciens HAB-2. BMC Microbio. 17 (2), 227 (2017). https://doi.org/10.1186/s12866-017-1134-z.

[3]Tsuge K., Ano T., Hirai M., et al. The Genes degQ, pps, and Ipa-8(sfp) Are Responsible for Conversion of Bacillus subtilis 168 to Plipastin Production. Antimicrobial Agents and Chemo. 43(9), 2183-2192 (1999). https://doi.org/10.1128/AAC.43.9.2183.