Project properties
Title Synthesis of a hybrid E.coli strain
Group Microbiology, Laboratory of
Project type thesis
Credits 12 BSc. thesis
Supervisor(s) Melvin Siliakus and Servé Kengen
Examiner(s) John van der Oost
Contact info melvin.siliakus@wur.nl
Begin date 2012/03/15
End date 2014/03/01
Description A main feature which distinguishes archaea from eukarya and bacteria are their membranes.
Whereas eukarya and bacteria have membranes composed of ester-lipids, archaea use ether- lipids, which lends them on exceptional robustness against extreme habitats. To further investigate the effect of isoprenoid ether lipids on membrane robustness, we plan to introduce the archaeal ether lipid biosynthesis pathway into a bacterium (E.coli). This requires the development of two synthetic operons comprising the MEP-DOXP (isoprenoïd precursor synthesis) pathway and an archaetidic acid synthesis pathway. Upon chromosomal integration the constructs will be induced chemically. In order to establish a partial or complete replacement of the ester-linked phospholipids, the down-regulation of the innate phosphatidate synthesis will be initiated simultaneously. This entails the exchange of the endogenous lysophosphatidic acid synthase promoter region with an inducible glucose/arabinose regulatory promoter. Additionally, the enzyme will be extended with a His-tag which allows an easy visualisation on western blot. Furthermore, the effect of this conditional knock-out will be characterized extensively.
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