User:Haddarr/sandbox

The T7 expression system is used in the field of microbiology to clone recombinant DNA using strains of E. coli. In order to perform this method, an expression vector is created which integrates two components: a T7 promoter and a gene of interest downstream of the promoter and under its control. The expression vector is transformed into one of several relevant strains of E. coli, such as BL21. The E. coli cell also has its own chromosome, which possesses a gene that is transcribed to transcribe T7 RNA polymerase. T7 RNA polymerase is responsible for beginning transcription at the T7 promoter of the transformed vector. The T7 gene is itself under the control of a lac promoter. Normally, both the lac promoter and the T7 promoter are repressed in the E. coli cell by the Lac repressor. In order to initiate transcription, an inducer must bind to the lac repressor and prevent it from inhibiting the gene expression of the T7 gene. Once this happens, the gene can be normally transcribed to produce T7 RNA polymerase. T7 RNA polymerase, in turn, can bind to the T7 promoter on the expression vector and begin transcribing its downstream gene of interest. To stimulate this process, IPTG can be added to the system. IPTG is a reagent which mimics the structure of allolactose, and can therefore bind to the lac repressor and prevent it from inhibiting gene expression. Once enough IPTG is added, the T7 gene is normally transcribed and so transcription of the gene of interest downstream of the T7 promoter also begins.