Difference between revisions of "Part:BBa K2971000"

Revision as of 21:02, 21 October 2019

crtE from Deinococcus radiodurans

This part is the gene crtE (dr1395) from the extremophile Deinococcus radiodurans and encodes geranylgeranyl diphosphate synthase (GGPPS). GGPPS catalyze the condensation of farnesyl pyrophosphate (FPP) and isopentenyl pyrophosphate (IPP) into geranylgeranyl diphosphate (GGPP)[1]. GGPP is a precursor for the carotenoid biosynthetic pathway. The sequence has been codon optimized for expression in Escherichia coli and iGEM registry compatibility.

The UiOslo 2019 team used this gene in a composite part (BBa_K2971004) to produce the red pigment lycopene, in Escherichia coli, by inserting it into a vector along with the genes crtB and crtI.

The part was cloned into an expression vector under the arabionose inducible promotor araC. Successful cloning was confirmed with colony PCR of transformed colonies (figure 1). Colonies that carried the insert was sequenced to check for the potential presence of mutations or frameshift.

We expressed the protein under the control of arabionose inducible promotor araC (figure 1). We detected a faint protein band during our SDS analysis; it is likely the expression was not well-optimized.

References 1. Liu, C., Sun, Z., Shen, S., Lin, L., Li, T., Tian, B., & Hua, Y. (2014). Identification and characterization of the geranylgeranyl diphosphate synthase in Deinococcus radiodurans. Letters in Applied Microbiology, 58(3), 219-224. doi:10.1111/lam.12181

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
COMPATIBLE WITH RFC[12]
21
INCOMPATIBLE WITH RFC[21]
Illegal BamHI site found at 841
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 43

@@ Line 12: / Line 12: @@
 <i>Escherichia coli</i>, by inserting it into a vector along with the genes <i>crtB</i> and <i>crtI</i>.
-We expressed the protein under the control of arabionose inducible promotor araC. We detected a faint protein band during our SDS analysis; it is likely the expression was not well-optimized.
+The part was cloned into an expression vector under the arabionose inducible promotor <i>araC</i>. Successful cloning was confirmed with colony PCR of transformed colonies (figure 1). Colonies that carried the insert was sequenced to check for the potential presence of mutations or frameshift.
+We expressed the protein under the control of arabionose inducible promotor araC (figure 1). We detected a faint protein band during our SDS analysis; it is likely the expression was not well-optimized.