Difference between revisions of "Part:BBa K1992003"
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==Experiments and results== | ==Experiments and results== | ||
| − | The fused Tar-GFP protein was cloned to our expression systems (<pratinfo>K1992008</partinfo> and <partinfo>K1992009</partinfo>) in order to examine the migration of the receptor. By using a fluorescence microscope it can be seen in figure 1 that the receptor is closter in the poles of the bacteria as expected. | + | The fused Tar-GFP protein was cloned to our expression systems ( <pratinfo>K1992008</partinfo> and <partinfo>K1992009</partinfo>) in order to examine the migration of the receptor. By using a fluorescence microscope it can be seen in figure 1 that the receptor is closter in the poles of the bacteria as expected. |
[[file:T--Technion_Israel--Tar_flourecent.png|600px|thumb|center|Fig1 (a) K1992004 - Tar expressed w RBS. (b) K1992005 - Tar expressed w the native RBS]] | [[file:T--Technion_Israel--Tar_flourecent.png|600px|thumb|center|Fig1 (a) K1992004 - Tar expressed w RBS. (b) K1992005 - Tar expressed w the native RBS]] | ||
Revision as of 14:16, 18 October 2016
Tar chemoreceptor tagged with GFP
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1282
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 2324
Illegal SapI.rc site found at 111
Usage and Biology
E.coli native chemoreceptors cluster in the cell poles. This property is critical for signal amplification and adaptation of the cell. Although little is known about the mechanism of localization, it is important to preserve this property with our designed receptors in order to keep a functional and sensitive chemotaxis response (1).
GFP labeling is a very common way to examine the migration and localization of certain proteins in vivo. Fusion of GFP to Tar chemoreceptor enabled us to track the migration and localization of the protein to the cell poles as expected. This part is a composed from three iGEM registry BioBricks.
Design considerations
The Tar receptor (BBa_K777000) and GFP(BBa_B0040) were taken and amplified from the kit. The flexible linker (J18921) was added by using reverse transcription PCR. The C-terminos of Tar
Experiments and results
The fused Tar-GFP protein was cloned to our expression systems ( <pratinfo>K1992008</partinfo> and BBa_K1992009) in order to examine the migration of the receptor. By using a fluorescence microscope it can be seen in figure 1 that the receptor is closter in the poles of the bacteria as expected.
Fig1. (a) Tar-GFP expression in UU1250 strain cloned with K1992008 expression system. (b) Tar expression in UU1250 strain cloned with K1992004 expression system.
Reference
1.SHIOMI, Daisuke, et al. Helical distribution of the bacterial chemoreceptor via colocalization with the Sec protein translocation machinery. Molecular microbiology, 2006, 60.4: 894-906.