Difference between revisions of "Part:BBa K2971004"

Revision as of 00:29, 22 October 2019

crtEBI under pBad promotor

This composite part contains the genes crtE, crtB and crtI combined into a single reading frame to produce lycopene when expressed in Escherichia coli. The genes express geranylgeranyl diphosphate synthase (GGPPS), phytoene synthase (CrtB) and phytoene desaturase (CrtI) respectively. GGPPS and CrtB produce the saturated compound phytoene, which is then desaturated by CrtI into the conjugated compound lycopene. When the part was expressed in E.coli, the pellet of the spun down expression culture acquired a pink color. The color validate that the part functions as intended.

Results

Figure 1: Expression of pBAD-crtEBI in Escherichia coli (BL21)
Expression was induced in 2% arabinose
From left to right :Uninduced E. coli with empty expression vector, Induced E.coli with empty expression vector, induced E.coli with crtEBI expression vector, uninduced E.coli with crtEBI expression vector.

The composite part was made by fusing crtEB (BBa_K2971010) and crtI (BBa_K2971001) via Gibson cloning. The part was then transformed into competent cells of E. coli. The presence of the insert was screened with colony pcr (figure 2).

Figure 2: Colony PCR of cells transformed with crtEBI
Only one colony (8) showed the presence of the correct insert. 8 was then further investigated.

Sequence and Features