Team:Tokyo Tech/Parts

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Parts Overview

To meet the criteria of the Gold Medal, we submitted BBa_K1949060
 and we characterized BBa_R0071, BBa_C0071, BBa_C0171, BBa_K1529300, BBa_K1529310, BBa_K1096001 and BBa_K1096002.

To meet the criteria of the Silver Medal, we submitted BBa_K1949050, BBa_K1949052, BBa_K1949030 and BBa_K1949032.

To meet the criteria of the Bronze Medal, we submitted BBa_K1949000, BBa_K1949001, BBa_K1949100, BBa_K1949101 and BBa_K1949102.

Favorite Tokyo Tech 2016 iGEM Team Parts

Name	Type	Description	Design	Length(bp)
BBa_K1949050	Coding	amiE	Yoshio Takata	1476
BBa_K1949060	Composite	Prhl(NM)-rbs-gfp	Yoshio Takata	799
BBa_K1949102	Composite	PBAD-rbs-mazF-tt-Ptet-rbs-gfp	Yoshio Takata	2520

Tokyo Tech 2016 iGEM Team Parts

Name	Type	Description	Design	Length(bp)
BBa_K1949000	Regulatory	Pcold	Yoshio Takata	313
BBa_K1949001	Measurement	Pcold-gfp	Yoshio Takata	1033
BBa_K1949020	Coding	yafN	Kazuki Fujisawa	297
BBa_K1949022	Composite	Plac-rbs-yafN	Kazuki Fujisawa	523
BBa_K1949030	Coding	yafO	Yoshio Takata	402
BBa_K1949031	Transrational unit	rbs-yafO	Yoshio Takata	420
BBa_K1949032	Composite	PBAD-rbs-yafO	Yoshio Takata	1638
BBa_K1949033	Composite	PBAD-rbs-yafO-tt-Ptet-rbs-gfp	Yoshio Takata	2580
BBa_K1949051	Transrational unit	rbs-amiE	Yoshio Takata	1494
BBa_K1949052	Composite	PBAD-rbs-amiE	Yoshio Takata	2712
BBa_K1949100	Composite	Plac-rbs-mazE	Yoshio Takata	565
BBa_K1949101	Composite	PBAD-rbs-mazF	Yoshio Takata	1575
BBa_K1949103	Composite	Ptet-rbs(BBa_B0034))-mazE	Yoshio Takata	332
BBa_K1949104	Composite	Ptet-rbs(BBa_J61117)-mazE	Yoshio Takata	332

1. Improved Part: BBa_K1949060

BBa_K1949060 meets the Gold Medal criteria!

We simulated our final genetic circuits and found that the circuits did not work, because Prhl activity was too weak compared to Plux. (see the Model page and the AHL Only Assay page). We therefore considered using the improved Prhl (BBa_K1529310, BBa_K1529300) established by iGEM 2014 Tokyo_Tech, but we noticed that they were inappropriate for two reasons (see the this page). Then, we decided to improve Prhl Promoter and obtain our original improved Prhl (Noticeable Mutant) (BBa_K1949060), hereafter referred to as Prhl(NM), that suited our goal.

Our purpose is to create strong Prhl for our final genetic circuits.

This experiment consists of the three parts below.

1. Improved Prhl by iGEM 2014 Tokyo_Tech and characterization of rhlR.




Fig. 4-1-1

Improvement of the wild type Prhl.




Fig. 4-1-2

Comparison of the improved Prhl by iGEM 2014 Tokyo_Tech to our original improved Prhl




Fig. 4-1-3

The past improved Prhl did not suit for our final circuits and we could construct the improved Prhl appropriate to our final circuits.

For more information, see this !

2.Best Basic Part: BBa_K1949050

BBa_K1949050 meets the Silver Medal criteria!

The Prince coli expresses AmiE protein, and Snow White coli recovers from its apparent death and wakes up again. We tested the function of AmiE protein that influences the end of the story.

Fig. 4-1-4

Our objective is to characterize the function of AmiE protein. We prepared three samples shown below. When we tested the AmiE degradation ability with these samples, the results show that C4HSL is not degraded by AmiE, but 3OC12HSL is degraded by AmiE.

• PBAD-rbs-amiE(pSB6A1)

• Ptet-rbs-luxR-tt-Plux-rbs-gfp (pSB6A1)

• Ptet-rbs-luxR-tt-Plux-rbs-gfp (pSB6A1)




Fig. 4-1-5

Best Composite Part: BBa_K1949102

BBa_K1949102 meets the Bronze Medal criteria!

The biggest attraction of the TA system is that it is able to control cell growth and synthesis of protein. In this experiment, mazE expression was induced by the addition of IPTG(2 mM) after MazF expression was induced by the addition of arabinose(0.02%). As a result, it was able to resuscitate from a state of being inhibited cell growth. We named this experiment as "Stop & Go" because it was to resuscitate growth from inhibiting cell growth.