Part:BBa_K4274036

PvanP*-patB_target

This is the composite part that we have designed to knock-out the patB gene in Bacillus subtilis. It starts with the promoter PvanP* (part: BBa_K4274006), a semi-synthetic promoter. Then, we have encoded the patB_target (part: BBa_ K4274007), the single guide RNA that locates the targeted gene sequence for Cas9 protein. This composite part terminates with the terminator B0015 (BBa_B0015). The parts used to knock-out patB from Bacillus subtilis, which is composed of patB_target (Part: BBa_K4274007), PvanP* (Part: BBa_K4274006).
This part collection was designed to knock-out patB gene in Bacillus subtilis, and then formed competition between Staphylococcus spp and modified Bacillus subtilis for the S-Cys-Gly-3M3SH but would not produce the malodorous molecule of 3M3SH.

Usage and Biology

We utilized the Cas9 protein to perform the knock-out of patB gene in Bacillussubtilis. patB_target (Part: BBa_K4274007) is a synthetic single guide RNA that assists the Cas9 protein to find the targeted gene sequence and perform the gene knock-out. As an extension, patB is one of the proteins in Staphylococcus. Spp. that breaks down S-Cys-Gly-3M3SH, resulting in the secretion of 3M3SH. Meanwhile, it was found that Bacillus subtilis has the identical gene sequence within its genome. Thus this part could be used to knock-out patB in B. subtilis to construct a promising bacterial strain for solving body odor problem.

Source

patB from Bacillus subtilis.

Characterization

Currently, our team succeed in knocking out the patB enzyme in the Bacillus subtilis 168, guarantee that Bacillus subtilis 168 can both absorb Cys-Gly-3M3SH and avoid the production of odor. This result had laid the experimental foundation and pave the way for our second-generation product.

Figure 1: Screening results of Bacillus subtilis strains with successful patB gene knock-out. (a) Electrophoresis results show that gene editing is successful. (b) The schematic representation of Bacillus subtilis genome before and after patB being knocked-out.

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]