Difference between revisions of "Part:BBa K1976052"

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<b>Figure 3</b>: 1% agarose gel electrophoresis, stained with HD-green.  
 
<b>Figure 3</b>: 1% agarose gel electrophoresis, stained with HD-green.  
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The protein product from this part was cleaned up and used in different concentrations to test its activity.
 
The protein product from this part was cleaned up and used in different concentrations to test its activity.
 
From left to right: pSB1C3 alone, pSB1C3 in the lysate buffer 1x, pSB1C3 in the lysate buffer 3x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976052 1x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976052 3x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976053 1x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976053 3x.
 
From left to right: pSB1C3 alone, pSB1C3 in the lysate buffer 1x, pSB1C3 in the lysate buffer 3x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976052 1x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976052 3x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976053 1x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976053 3x.
 
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Revision as of 11:26, 27 October 2016

Colicin E2 DNase domain and immunity protein Im2 generator with anderson promoters

Generator of colicin E2 DNase domain BBa_K1976048 and immunity protein Im2 BBa_K1976026.

Figure 1: The structure of the Im2 immunity protein and Colicin E2 in a complex.


Usage

This part can be used to produce colicin without killing the host cell, as the immunity protein is produced more than equimolar to the DNase fragment. If BBa_K1976028 is used instead of BBa_K1976027, it can function as a amber-suppression inducible killswitch.

Characteristics


Figure 2: SDS-PAGE of the Colicin E2 DNase domain, the blue arrow marks the band of the expression. From left to right: Top10 E. coli, Top10 E. coli transformed with BBa_K1976052 and Top10 E. coli transformed with BBa_K1976053.


Figure 3: 1% agarose gel electrophoresis, stained with HD-green. The protein product from this part was cleaned up and used in different concentrations to test its activity. From left to right: pSB1C3 alone, pSB1C3 in the lysate buffer 1x, pSB1C3 in the lysate buffer 3x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976052 1x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976052 3x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976053 1x, pSB1C3 incubated with lysate of Top10 transformed with BBa_K1976053 3x.


Molecular Weight Colicin E2 Dnase domain
15.2 kDa
Immunity Protein
10.0 kDa
Residues Colicin E2 Dnase domain 133
pI Colicin E2 Dnase domain
9.06
Immunity Protein
4.66
[Data taken created in Snapgene and http://isoelectric.ovh.org/calculate.php]

Characteristics

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
    Illegal NheI site found at 477
    Illegal NheI site found at 500
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 681
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 89
  • 1000
    COMPATIBLE WITH RFC[1000]


[1] Cascales et al, Colicin Biology, Microbiology and Molecular Biology Reviews, vol. 71, pp. 158-229, 2007