Part:BBa_K3440016

Mtrb under constitutive promoter

Pconst(BBa_J23100) - RBS(BBa_B0034) - Mtrb(BBa_K3146001) - Myc(BBa_K823036)

Usage and Biology

This part expresses mtrb constitutively. Mtrb is responsible for electricity production in Shewanella oneidensis (UniProtKB - Q8CVD4). In our project, we created this part as an intermediate to build BBa_K3440019 and BBa_K3440021.

Characterization

Due to the pandemics, we haven’t been able to use biobricks to create the iGEM Stockholm 2020 parts. Those parts were ordered as gene blocks from Integrated DNA Technologies Inc..

After heat shock transformation of the pSB1C3 plasmid containing the BBa_K3440016, we picked colonies from plates and PCR amplified them with primers VF and VR2.

We ran gels of the product at 180V and for 30 mins (Figure 2). We obtained the expected size for the bands (2499bp) for cf3.

Figure 2: Colony PCR Gel for BBa_K3440016 (cf)

We therefore prepared plasmid preparations and glycerol stocks of those and sent some of them for sequencing to Microsynth AG. The sequence obtained did not correspond to mtrb. The part had not been insert into the plasmid. We therefore highly recommend other teams to always send their samples for sequencing, even when the bands on gels have the right size

No further characterisation was done since this was only an intermediate part used to build others.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 2184
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]