Difference between revisions of "Part:BBa K3182108"

Revision as of 07:16, 20 July 2019

pT7-CBDcipA-sfGFP

A cellulose binding domain (CBDcipA) from Clostridium thermocellum Cellulose scaffolding protein (CipA) which can be used to purify or attach proteins to cellulose. The part also has a flexible GS-linker (-GGGGSGGGGS-) with a thrombin site (-LVPRGS-, thrombin RS) added at the end, clevage with thrombin will add one glycine and one serine to the N-terminal of the C-terminal fusion protein of the CBDcipA. Also added a BamHI recognition sequence (BamHI RS) to enable changeable fusion protein to the CBDcipA. BamHI was chosen because its RS codes for one glycine and one serine, fitting it to the end of the thrombin site, BamHI is also a cheap enzyme and can be used with methylated DNA.

The part has a very strong expression with a T7 promotor (BBa_I719005) as well as a 5'-UTR (BBa_K1758100) region which has been shown to further increase expression in E. coli (BBa_K1758106), ([http://www.ncbi.nlm.nih.gov/pubmed/2676996 Olins et al. 1989]), ([http://www.ncbi.nlm.nih.gov/pubmed/23927491 Takahashi et al. 2013]). Both this part and the part were sfGFP was changed for AsPink (BBa_K3182000) showed great expression.

Cellulose binding domain from Clostridium thermocellum fused to sfGFP to further characterize the CBD and linker construct. Figure 1 Figure text here Figure 2 Figure text here Figure 3 Figure text here

Figure 1 Figure text here

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 580
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
INCOMPATIBLE WITH RFC[1000]
Illegal SapI.rc site found at 598

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 A cellulose binding domain (CBDcipA) from Clostridium thermocellum Cellulose scaffolding protein (CipA) which can be used to purify or attach proteins to cellulose. The part also has a flexible GS-linker (-GGGGSGGGGS-) with a thrombin site (-LVPRGS-, thrombin RS) added at the end, clevage with thrombin will add one glycine and one serine to the N-terminal of the C-terminal fusion protein of the CBDcipA. Also added a BamHI recognition sequence (BamHI RS) to enable changeable fusion protein to the CBDcipA. BamHI was chosen because its RS codes for one glycine and one serine, fitting it to the end of the thrombin site, BamHI is also a cheap enzyme and can be used with methylated DNA.
-The part has a very strong expression with a T7 promotor (BBa_I719005) as well as a 5'-UTR and a g10-L RBS (BBa_K1758100) region which has been shown to further increase expression in E. coli (BBa_K1758106), (Olins et al. 1989), (Takahashi et al. 2013). Both this part and the part were sfGFP was changed for AsPink (BBa_K3182000) showed great expression.
+The part has a very strong expression with a T7 promotor (<partinfo>BBa_I719005</partinfo>) as well as a 5'-UTR (<partinfo>BBa_K1758100</partinfo>) region which has been shown to further increase expression in E. coli (<partinfo>BBa_K1758106</partinfo>), ([http://www.ncbi.nlm.nih.gov/pubmed/2676996 Olins et al. 1989]), ([http://www.ncbi.nlm.nih.gov/pubmed/23927491 Takahashi et al. 2013]). Both this part and the part were sfGFP was changed for AsPink (<partinfo>BBa_K3182000</partinfo>) showed great expression.
 Cellulose binding domain from Clostridium thermocellum fused to sfGFP to further characterize the CBD and linker construct.