Difference between revisions of "Part:BBa K4239003:Design"
(→‎Design Notes)
 
(One intermediate revision by the same user not shown)
Line 10: Line 10:
 
<html>
 
<html>
  
−
<p><i>fiatluxA</i> did not have any mutation compare to <i>iluxA</i>, because no igem restriction site (EcoR1, Xba1, Spe1 and Pst1) were into the gene.
+
<p><i>fiatluxA</i> did not have any mutation compare to <i>iluxA</i>, because no igem restriction site (EcoRI, XbaI, SpeI and PstI) were into the gene.
  
 
<p><i>iluxA</i> had mutations from <i>luxA</i> (K22E, T119A, S178A). They changed the protein sequence, in order to create a system that produced more light. The first and the last letters correspond to the Amino Acid before and after mutation, and the number indicates its position in the protein. These mutations were error-prone PCR induced according to Gregor et al.’s study in 2018.</p>
 
<p><i>iluxA</i> had mutations from <i>luxA</i> (K22E, T119A, S178A). They changed the protein sequence, in order to create a system that produced more light. The first and the last letters correspond to the Amino Acid before and after mutation, and the number indicates its position in the protein. These mutations were error-prone PCR induced according to Gregor et al.’s study in 2018.</p>
Line 20: Line 20:
 
<html>
 
<html>
  
−
The source of <i>fiatluxA</i> was the <i>ilux</i> operon available in a pGEX plasmid. An overlap PCR was performed to reconstitute directly <i>fiatluxABE</i> fragments which had been cut by the restriction enzymes.  
+
<p>The source of <i>fiatluxA</i> was the <i>ilux</i> operon available in a pGEX plasmid. An overlap PCR was performed to reconstitute directly <i>fiatluxABE</i> fragments which had been cut by the restriction enzymes.</p>
  
 
</html>
 
</html>

Latest revision as of 00:12, 12 October 2022


Enhanced luciferase substrate forming units fiatluxA


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 504
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 1023


Design Notes

fiatluxA did not have any mutation compare to iluxA, because no igem restriction site (EcoRI, XbaI, SpeI and PstI) were into the gene.

iluxA had mutations from luxA (K22E, T119A, S178A). They changed the protein sequence, in order to create a system that produced more light. The first and the last letters correspond to the Amino Acid before and after mutation, and the number indicates its position in the protein. These mutations were error-prone PCR induced according to Gregor et al.’s study in 2018.

Source

The source of fiatluxA was the ilux operon available in a pGEX plasmid. An overlap PCR was performed to reconstitute directly fiatluxABE fragments which had been cut by the restriction enzymes.

References

Gregor C, Gwosch KC, Sahl SJ, Hell SW. Strongly enhanced bacterial bioluminescence with the ilux operon for single-cell imaging. Proc Natl Acad Sci U S A. 2018 Jan 30;115(5):962-967. doi: 10.1073/pnas.1715946115. Epub 2018 Jan 16. PMID: 29339494; PMCID: PMC5798359.