Fig. 5.

Comparison of growth, gene transcription, translation and isobutanol production in engineered strains Syn-pEEK2, pEEK2-kivd, Syn-IB-7, -8, -9 and -6. A) RT-PCR, SDS-PAGE and Strep-tag Western-immunoblot (top to bottom). Each lane represents the results from a single engineered strain. –RT is a negative RT-PCR control using RNA as template without addition of RT enzyme to control for possible DNA contamination. The positive control is an RT-PCR carried out using the corresponding plasmid as template. Red arrows indicate the location of kivd, blue arrows indicate the expected location for different AHDs. The first row of the Strep-tag Western-immunoblot shows the expression of kivd and the second row shows the expression of all the ADHs. The negative control for protein gel and Western-immunoblot is the empty vector strain Syn-pEEK2. B) Growth curve during 7 days of cultivation. C) Isobutanol production at day 3, day 5 and day 6 from all the engineered strains. Results are the mean of 4 biological replicates and 3 technical replicates, error bars represent standard deviation. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)