Part:BBa_K1111013
Ice Nucleation Protein fused to Streptavidin Dead
Introduction
INP sequence fused with Streptavidin Dead by gibson assembly. The INP-streptavidin construct expresses streptavidin at the outer membrane of E.Coli. INP is an outer membrane protein foud in the genome of Pseudomonas syringae. It is under the control of Lac promoter and thus is constitutively expressed in E.Coli. The expression of this plasmid was tested by transforming cells and looking for microscopy at either of a fluorescent antibody again streptavidin or a fluorescent biotin (biotin and streptavidin have a strong affinity).
Usage and Biology
We used this part as a negative control. We also have two other parts implying streptavidin non mutated, which are supposed to bien biotin (BBa_K1111012 and BBa_K1111014), and this streptavidin is mutated to decrease the Kd of the binding streptavidin-biotin. Thus, the streptavidin dead, although it is well exported to outer membrane, won't bind to biotin, or will bind weakly.
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal XbaI site found at 1727
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23INCOMPATIBLE WITH RFC[23]Illegal XbaI site found at 1727
- 25INCOMPATIBLE WITH RFC[25]Illegal XbaI site found at 1727
Illegal NgoMIV site found at 1036
Illegal AgeI site found at 1691
Illegal AgeI site found at 1742 - 1000COMPATIBLE WITH RFC[1000]
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