Team:Gunma/Parts

Team:Gunma Parts

Parts

Parts Overview

Firstly we made a basic part of magainin-2 sequense, toxic protein that derives from Xenopus laevis. Secondly, we made its expression visible by making a composite part that expresses GFP with magainin-2.

BBa_K3050001

BBa_K3050000

Initially we were strongly inspired by the project of UT-Tokyo2016,color-changing E.coli dependently on cell divisions and determined to develop their project. Our current project is kind of a section of the initial project which was too content-full for a year.

Our Initial Project

Generation 1 RED

Generation 2 BLUE

Generation 3 GREEN

+

Generation 4 DEATH!!

Designed by team UT-Tokyo 2016, cited from

https://2016.igem.org/Team:UT-Tokyo

1. BBa_K3050000 (Partially validated)

This is a composite part that includes T7 promotor + RBS + mag-2 + GFP + double terminators.

BBa_I746909 is linerized by Inverse PCR to be combined with magainin-2 sequense by In-Fusion reaction.

BL21(DE3) transformed pSB1C3 BBa_K3050000 successfully grew and emit strong green fluorescent light as shown in pictures.

Thus we succeeded to prove Inverse PCR,In-Fusion reaction,and transformation went great.

GFP part is on downstream of T7 promotor and magainin-2, hence we're quite sure that this result indicates that magainin-2 is expressed at the same time.

Fig.1 BL21(DE3) pSB1C3 BBa_K3050000 expressed under visible light with negative control without IPTG induction

BL21(DE3) pSB1C3 BBa_K3050000, under visible light, 3.5 hours after directly added 300 μM IPTG.

Fig.2 pSB1C3 BBa_K3050000 expressed under UV light with negative control without IPTG induction

BL21(DE3) pSB1C3 BBa_K3050000, under UV light, 3.5 hours after directly added 300 μM IPTG.

Fig.3 Four biological replicates all highly expressed

BL21(DE3) pSB1C3 BBa_K3050000, under visible light, 26 hours after directly added 500 μM IPTG.

The fluorescent light didn't extingished for more than whole a day after added 500 μM IPTG.

2. BBa_K3050001 (Validated as a part of BBa_K3050000)

Part Table

iGEM19 Gunma

Name	Type	Description	Designer	Length
BBa_K3050000	Translational_Unit	pT7+RBS+mag-2+GFP+double_terminator	Kita Mizuki	1171
BBa_K3050001	Coding	magainin-2_weaken_cell_membrane	Kita Mizuki	72