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Guide RNA (gRNA) is used as a molecule that helps with the cleavage of DNA since it guides the Cas nuclease to a targeted dsDNA sequence.  As a gRNA, sfp_target is used for knocking-out the natural, invalid sfp gene, and then knocking-in sfp (Part number:BBa_K4274009) and degQ(Part number:BBa_K4274010) gene in the genome of <i>Bacillus subtilis</i>.  
 
Guide RNA (gRNA) is used as a molecule that helps with the cleavage of DNA since it guides the Cas nuclease to a targeted dsDNA sequence.  As a gRNA, sfp_target is used for knocking-out the natural, invalid sfp gene, and then knocking-in sfp (Part number:BBa_K4274009) and degQ(Part number:BBa_K4274010) gene in the genome of <i>Bacillus subtilis</i>.  

Latest revision as of 05:09, 12 October 2022

sfp_target(gRNA)

Guide RNA (gRNA) is used as a molecule that helps with the cleavage of DNA since it guides the Cas nuclease to a targeted dsDNA sequence. As a gRNA, sfp_target is used for knocking-out the natural, invalid sfp gene, and then knocking-in sfp (Part number:BBa_K4274009) and degQ(Part number:BBa_K4274010) gene in the genome of Bacillus subtilis.

Usage and Biology

Single-guide RNA (sgRNA) is RNA designed artifially to be able to bind to a DNA sequence. It combines with the Cas9 protein to work to cleave the target RNA and though it is slightly shorter, it has the same function as the original RNAs in the CRISPR/Cas9 system. We designed the sfp_target(gRNA) to target the region of the natural sfp gene to knock-out and simultaneously knock-in sfp (Part number:BBa_K4274009) and degQ (Part number:BBa_K4274010) gene in situ. After the sucessful modification, Bacillus subtilis 168 was allowed to produce fengycins. This could be reused by other teams to edit the genome of Bacillus subtilis and thus realize fengycins’ production.

Source

Bacillus subtilis

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Reference

[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.