Difference between revisions of "Part:BBa K3050000"

Latest revision as of 23:28, 21 October 2019

pT7+RBS+mag-2+GFP+double_terminator

Magainin-2 expressed inside E.coli combines to cell membrane and damages it. In our project, we aimed to make BL21(DE3) lyse easier by transforming BBa_K3050000 on pSB1C3.

Usage and Biology

This is a composite part that includes T7 promotor + RBS + mag-2 + GFP + double terminators. You can use this part when you want to weaken cell membrane of E.coli,also you can visually know when it expressed. As the magainin-2 and GFP is under T7 promotor, you can easily induce it by adding IPTG. We recommend 300 μM IPTG for directly putting on colonies on plates, and 500 μM for culture solution. You want to know more what is it like when IPTG is added? Read this page more or see also: https://parts.igem.org/Part:BBa_I746909:Experience

BBa_I746909 is linerized by Inverse PCR to be combined with magainin-2 sequense by In-Fusion reaction. https://parts.igem.org/Part:BBa_I746909

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

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

Fig.3 Four biological replicates all highly expressed

Fig.4 500 μM IPTG induced BBa K3050000, strongly emitting green fluorescent light

Sequence and Features