In vivo study of gene regulation and gene function in Sulfolobus ssp.: application and development of transformation vectors

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Sulfolobus represents one of the best studied hyperthermophilic archaea of the kingdom crenarchaeota and has developed into an important model organism in particular for the understanding of information processing (i.e. replication, transcription, translation) in archaea. However, no efficient genetic system is yet available for Sulfolobus and for other hyperthermophiles. Recent efforts in our laboratories including the characterization of a Sulfolobus plasmid and the development of a stable and selectable vector system have opened up new avenues to study gene regulation and protein/protein interaction in vivo and to refine and to develop genetic tools. We will use our novel vector system for the characterization of regulated promoters in vivo, and for the expression of tagged proteins. Furthermore, we will refine the transformation system such that it will allow high efficient cloning in Sulfolobus and we want to develop novel tools for mutagenesis and/or gene silencing that will allow the straight forward study of gene functions in a hyperthermophilic archaeon. The proposed approaches include the development of novel markers and DNA uptake techniques with liposomes, a helper phage system, transposon mutagenesis and gene silencing by homologous recombination or RNA interference.