Part:BBa_K2213006

LowPromoter_PduD(1-20)_mCherry

This part is an improved version of https://parts.igem.org/Part:BBa_K562001, submitted by Dundee 2011.

The PduD(1:20) tag is one of many tags that has been proved to localise inside a Pdu microcompartment. This has been proved by the iGEM Dundee 2011 team http://2011.igem.org/Team:Dundee. This part has been tagged with the fluorescent protein, mCherry, to facilitate its characterisation, as can be seen in the figure below:

figure1. circuit diagram of BBa_K2213006.

Improvements

This part is an improved version of https://parts.igem.org/Part:BBa_K562001, originally submitted by team Dundee 2011. The original part contained an illegal XbaI site which has been removed to make it biobrick compatible.
This part has also been expressed under different strength promoters, by Manchester2017, to find the optimal level of induction.

• A low strength Anderson promoter (here)
• medium strength Anderson promoter (https://parts.igem.org/Part:BBa_K2213007)
• high strength Anderson promoter (https://parts.igem.org/Part:BBa_K2213008)

Characterisation

The PduD tag was combined with the low strength Anderson promoter (https://parts.igem.org/Part:BBa_J23105) and mCherry.

A gradient of fluorescence is evident when compared to medium and high promoters.




The expression levels compared to medium and high are as shown.





When expressed alone the distribution of the mCherry signal was homogeneous and fluorescence level was very low making visualisation difficult. In the presence of EutSMNLK the mCherry signal was much brighter and clumped together, indicating localisation to the BMC.

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 8
  Illegal NheI site found at 31
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]