Abstract
The fission yeast Schizosaccharomyces pombe is an excellent genetically tractable model organism used in the study of conserved eukaryotic cellular biology. One genetic tool in the assessment of gene function is the in vivo overexpression of proteins. Existing overexpression tools have limitations of induction kinetics, dynamic range, and/or system-wide changes due to the induction conditions or inducer. Here, I describe the methodology for the use of a plasmid-based long non-coding RNA (lncRNA)-regulated overexpression system that is induced by the addition of thiamine. This system, termed the pTIN-system (thiamine inducible), utilizes the fast repression kinetics of the thiamine-regulated nmt1 + promoter integrated with the lncRNA regulated tgp1 + promoter. The advantages of the pTIN-system are rapid induction kinetics of gene expression, broad dynamic range, and tunable expression.