Difference between revisions of "Part:BBa K2669001:Design"

Latest revision as of 10:51, 27 September 2018

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

Design Notes

To make this part, we had to modify Part:BBa_K2003011 by moving the start codon to the front of the histidine tag, as opposed to after it. We also wished to express this protein constitutively as opposed to inducibly in order to make it more convenient for us to test if our part worked. Therefore we created the composite Part:BBa_K2669000 in order to express this part and conduct tests to verify that it works.

In order to prevent potential protein aggregation and/or strain on the cells we expressed our part in a low copy ampicillin backbone ordered from IDT (pUCIDT). In addition, VR and VF2 cloning sites were inserted into our part for easy sequencing/PCR.

We also further modified Part:BBa_K2003011 by adding a stop codon (taa) at the end of the flexible GSG linker in order to use it in our composite Part:BBa_K2669000. It is very important to note that this stop codon should be removed if one wishes to use this part to create a fusion protein.

References

Kumagai, A., Ando, R., Miyatake, H., Greimel, P., Kobayashi, T., Hirabayashi, Y., Shimogori, T., and Miyawaki, A. (2013).
A Bilirubin-Inducible Fluorescent Protein from Eel Muscle. Cell 153, 1602–1611.

@@ Line 4: / Line 4: @@
 <partinfo>BBa_K2669001 SequenceAndFeatures</partinfo>
+===Design Notes===
+To make this part, we had to modify [[Part:BBa_K2003011]] by moving the start codon to the front of the histidine tag, as opposed to after it.  We also wished to express this protein constitutively as opposed to inducibly in order to make it more convenient for us to test if our part worked.  Therefore we created the composite [[Part:BBa_K2669000]] in order to express this part and conduct tests to verify that it works.
+In order to prevent potential protein aggregation and/or strain on the cells we expressed our part in a low copy ampicillin backbone ordered from IDT (pUCIDT).  In addition, VR and VF2 cloning sites were inserted into our part for easy sequencing/PCR.
+We also further modified [[Part:BBa_K2003011]] by adding a stop codon (taa) at the end of the flexible GSG linker in order to use it in our composite [[Part:BBa_K2669000]].  It is very important to note that this stop codon should be removed if one wishes to use this part to create a fusion protein.
 ===References===