Part:BBa_J23117:Experience

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Applications of BBa_J23117

Evaluation of Anderson promoter J23117 in B. subtilis by iGEM-Team LMU-Munich 2012

This Anderson promoter was evaluated without fused RFP with the lux operon as a reporter in B. subtilis. See the new BioBrick BBa_K823013 without RFP and have a look at the [http://2012.igem.org/Team:LMU-Munich/Data/Anderson Data] from the evaluation in B. subtilis.

User Reviews

UNIQ3ab12307daa38171-partinfo-00000000-QINU UNIQ3ab12307daa38171-partinfo-00000001-QINU

••••• iGEM Team Göttingen 2013

Additionally to our characterization of this part, we also used it for our reporter system and it worked very good! We also improved it by switching the pre- and suffix, basically inverting it. This way, we were able to use it in an "inverted" expression unit on the same vector as our reporter system. For further information see: https://parts.igem.org/wiki/index.php?title=Part:BBa_K1045011

UNIQ3ab12307daa38171-partinfo-00000003-QINU

••••• iGEM15_UNITN-Trento

We used the part BBa_J23117 in the InterLab Measurement context. We assembled a measurement device by inserting the part BBa_I13504 amplified by PCR into the part BBa_J23117. The device was characterized in E. coli NEB10β, JM109, and NEB Express by measuring GFP expression with a cuvette-based spectrofluorometer, a fluorescence plate reader, and a flow cytometer. We also confirmed the promoter activity with a cell-free S30 extract system and measured mRNA by RT-qPCR. For a better understanding on protocols and characterizations, please check out our Wiki page UNITN-Trento iGEM 2015!

Evaluation of Anderson promoter J23117 in E. coli by [http://2013.igem.org/Team:Goettingen iGEM Göttingen 2013]

Shown here:
Upper two pictures: Growth curves of promoter strains on the left, growth curves of control strains on the right. Three biological replicates are shown.
Middle two pictures: RFP/OD600 of promoter strains on the left, RFP/OD600 of control strains on the right. Three biological replicates are shown.
Bottom three pictures: qRT PCR promoter analyses in three different growth phases. Promoters are normalised against BBa_J23117 .

Promotor 1:BBa_J23117
Promoter 2:BBa_J23116
Promoter 3:BBa_J23110
Promoter 4:BBa_J23118

The promoter strength was measured by using the reporter gene rfp.
Three different approaches were used: 1. RFP measurement, 2. qRT-PCR analyses and 3. single cell microscopy. Moreover, the first and the second approach characterised the promoter activity along the growth curve and to three important time points, respectively.
Taken together, the majority of our results from these approaches showed that BBa_J23117 has the lowest promoter activity compared to BBa_J23116, BBa_J23110 and BBa_J23118. For more details, visit the discussion on our wikipage.

Fig. 1. RFP and qRT-PCR promoter analyses.

Fig. 2. Microscopic promoter analyses on single cell level
on the left side: bright field (BR), on the right side: RFP filter (rfp)
P1: BBa_J23117, P2: BBa_J23116, P3: BBa_J23110, P4: BBa_J23118, P8: control

••••• University of Texas at Austin iGEM 2019

Characterization of the Anderson series through burden monitoring by the University of Texas at Austin's 2019 iGEM team Description Our team transformed the Anderson series of RFP reporters (J23101, J23113, J23104, J23107, J23117 in pSBC13 backbone) into our constitutive GFP burden monitor E. coli strain and measured the relative burden of each part. They contained a constitutive GFP sequence in the genome which serves as a way to measure the constructs’ ribosome allocation. Our results show a certain reduction ingrowth rate for each part as a result of ribosome misallocation away from the genome and towards the plasmid containing the construct. The promoter strengths associated with each RFP reporter construct shows that parts with stronger promoters express less GFP and have a reduced growth rate when compared to the constructs containing weaker promoters. The promoters associated with these RFP reporters were used to create a series of BFP reporters, also transformed into our GFP burden monitor strain, and create the regression on the figure below. For more information on characterization of these parts through burden monitoring and evolutionary stability experiments, visit our team’s wiki page: [1]