Regulatory

Part:BBa_K3855004

Designed by: Sebastian Bayer   Group: iGEM21_BOKU-Vienna   (2021-09-25)


PrRNA42- weak lactobacilli promotor

A weak constitutive Promotor for Lactobacillus plantarum. GusA activity of approximately 350 MU.


We confirmed the activity using mCherry as signalling protein. Simultaneously, we compared it to 7 other promotors. Five of those we added to the registry and where published by Rud et al 2006 . Two were published by Team Austin, Texas, 2017.

Experimental set up

5ml cultures of the eight different transfected Lactobacillus plantarum NC8, as well as an untransfected control, were incubated at 30°C overnight under anaerobic conditions and constant shaking. On the following day, conditions were changed to aerobic for 3-4 hours. Afterwards, the cell suspensions were diluted to different densities (OD 600 of 2, 1 and 0.1). The measurement was carried out using a Tecan reader. Samples where excited at 587 nm and emission measured at 640 nm. Sterile media was used as a reference. Experiments were carried out in triplets.

Construct Promotor
1 BBa_K3855000
2 BBA_K3855001
3 BBa_K3855002
4 BBA_K3855003
5 BBA_K3855004
6 BBA_K3855005
7 BBa_K2253000
8 BBa_K2253001


We found that the promotor expression is of low strength.

Figure 1. Excitation at OD 2,1 and 0.1 of different mCherry transfected L.plantarum suspensions. Wildtype emission was subtracted. Raw values can be found in our Benchling write up (Cloning, Friday 8.10.2021).


Rud et al. 2006: A synthetic promoter library for constitutive gene expression in Lactobacillus plantarum MICROBIOLOGY Volume 152, Issue 4.

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]


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Categories
Parameters
None