Coding
Part:BBa_K4361317
Designed by: Lars van den Biggelaar Group: iGEM22_TUDelft (2022-09-19)
BlcR V68K
A mutant of the BlcR protein, created through site-directed mutagenesis with primers R3 (Part:BBa_K4361218) and F3.8 V68K (Part:BBa_K4361226). For this mutant, the valine in position 68 has been changed to lysine by mutating the GTG codon to AAA.
This mutant also contains the following nucleotide mutations outside of the targeted site:
- C 497 > T, resulting in substitution S166L
- G 628 > A, resulting in substitution D210N
Sequence and Features
Assembly Compatibility:
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 694
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 78
- 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI.rc site found at 589
Usage and Biology
The set of BlcR mutants (Part:BBa_K4361300 through Part:BBa_K4361319) were expressed in a PURE system in the presence of GreenLys (fluorescent Cy2-labeled lysine). Samples were then run on an SDS PAGE gel (Figure 1.) to determine whether or not the protein was produced. For this mutant (lane 18), a protein band corresponding to the BlcR monomer is visible. Concluding that the production in an E. coli cell-free system was successful for this mutant.
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Categories
Parameters
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