Difference between revisions of "Part:BBa E0040:Experience"

USTC_China iGEM13

In USTC_China iGEM13's project,we bulit the N-terminal TD1 modified GFP to make it is capable of transporting through skin barrier, then induced expression in BL21 and Bacillus Subtilis WB800N with IPTG and tested positive by SDS-PAGE and ultraviolet ultraviolet spectrometry. Students in Wen lab validated the efficient transdermal function of N-terminal TD1 modified GFP(not BBa_E0040 ).

figure 1 SDS-PAGE of N-terminal TD1 modified GFP

figure 2 Fluorescence detection of N-terminal TD1 modified GFP expression with and without IPTG induction

figure 3 Our Transdermal test device

figure 4 Results of transdermal test of N-terminal TD1 modified GFP and unmodified GFP

Reference

SUN Zheng, WEN Long-ping.Transdermal delivery of fusion green fluorescent protein mediated by covalently associated TD1 peptide. OURNAL OF UNIVERSITY OF SCIENCE AND TECHNOLOGY OF CHINA.2009.Vol.39,No.4

KAIST_iGEM_2012

Figure 1. E.coli strain MG1655 expressing BBa_E0040 under control of BBa_K907004 after overnight culture. 3mL culture with M9 media in 14 mL round bottom tube(Top left two) and centrifuged cells in eppendorf tube(Bottom left two). The expression of BBa_0040 is clearly observed with naked eye after overnight culture.

BBa_E0040 was successfully used to produce GFPmut3b in E.coli strain MG1655 in LB or M9 minimal media under the control of promoter-BBa_J23119 and RBS-BBa_B0034 in the Dual Phase Protein Generator(GFP default), BBa_K907004.

Antiquity

This review comes from the old result system and indicates that this part worked in some test.

iGEM11_WHU_China

This part was used by WHU_China team as a reporter of long-term oscillator. Information from the website of this part shows that its length is about 720bp; however, the result of restriction enzymes cleavage experiment showed that it is actually about 1kb. And after several attempts, we failed to assemble this part with other biobricks.

iGEM Dundee 2013

The function of this part was characterised and quantified in a new salt-responsive device. The GFP encoded by BBa_E0040 was hooked up to an OmpR-responsive promoter (BBa_R0083) via a ribosome binding site (BBa_B0034) to produce a new composite part (a reporter) (BBa_K1012005). GFP fluorescence could be detected in an E. coli chassis, and we used an envZ mutant strain to show the sensor was working correctly.

iGEM Brasil-USP 2015

Designed by Brasil-USP team, BBa_K1819000 is a GFP coding sequence with an N-terminal linker, which is 5’-flanked by NdeI restriction site.
It represents an improvement from BBa_E0040 due to addition of NdeI restriction site, allowing other IGEM teams to express a chimera C-terminally attached to GFP, which exhibits bright green fluorescence when exposed to light in the blue to ultraviolet range.
NdeI restriction site avoids any additional amino acid residue(s) between coding sequence and the fused N-terminal tag, ideal for recombinant protein expression.