Difference between revisions of "Part:BBa K3182100"
Line 5: Line 5:
 
AsPink dropout enabling colour-screening for positive colonies. Using BamHI and SpeI or PstI on both this part assembled in pSB1C3 and the insert of choice will yield a fusion protein between CBDcipA and the insert. The fusion protein can later be cleaved with thrombin to yield two separate proteins. The C-terminal fusion will have one glycine and one serine added to the N-terminal of the protein.
 
AsPink dropout enabling colour-screening for positive colonies. Using BamHI and SpeI or PstI on both this part assembled in pSB1C3 and the insert of choice will yield a fusion protein between CBDcipA and the insert. The fusion protein can later be cleaved with thrombin to yield two separate proteins. The C-terminal fusion will have one glycine and one serine added to the N-terminal of the protein.
  
−
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]). This requires the T7-RNA-polymerase and bacteria with this should only be used after the C-terminal fusion protein has been assembled. Systems with the T7-RNA-polymerase can be used as well for assembly of CBD-fusion proteins, but will also express the CBD part and until it reaches a stop codon in the constitutive promoter.  
+
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]). This requires the T7-RNA-polymerase and bacteria with this should only be used after the C-terminal fusion protein has been assembled.  
  
 
[[File:T--Linkoping_Sweden--pinkwhite.png|800px|thumb|center|<b>Figure 1.</b> ]]
 
[[File:T--Linkoping_Sweden--pinkwhite.png|800px|thumb|center|<b>Figure 1.</b> ]]

Revision as of 10:32, 15 July 2019


pT7-CBDcipA-pCons-AsPink

AsPink dropout enabling colour-screening for positive colonies. Using BamHI and SpeI or PstI on both this part assembled in pSB1C3 and the insert of choice will yield a fusion protein between CBDcipA and the insert. The fusion protein can later be cleaved with thrombin to yield two separate proteins. The C-terminal fusion will have one glycine and one serine added to the N-terminal of the protein.

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]). This requires the T7-RNA-polymerase and bacteria with this should only be used after the C-terminal fusion protein has been assembled.

Figure 1.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 592
    Illegal NheI site found at 615
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 580
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]