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Isopropyl β-d-1-thiogalactopyranoside (IPTG) is a molecular biology reagent. This compound is a molecular mimic of allolactose , a lactose metabolite that triggers transcription of the lac operon , and it is therefore used to induce protein expression where the gene is under the control of the lac operator .
In E. coli BL21(DE3) the expression of the T7-RNAP is suppressed by the constitutively expressed LacI repressor. LacI binds the lac operator, which is located downstream of the LacUV5 promoter, preventing the production of the T7-RNAP. However, upon supplementation of IPTG, the LacI repressor dissociates from the lac operator, allowing for the ...
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. [6]
Additionally, unlike the lac promoter, lacUV5 works independently of activator proteins or other cis regulatory elements (apart from the -10 and -35 promoter regions). [2] While no activators are required, lacUV5 promoter expression can be regulated by the LacI repressor and can be induced with IPTG , which is an effective inducer of protein ...
IPTG, a molecule similar to lactose, but with a sulfur bond that is not hydrolyzable so that E. coli does not digest it, is used to activate or "induce" the production of the new protein. Once the cells are induced, it is difficult to remove IPTG from the cells and therefore it is difficult to stop expression.
Its production may be induced by a non-hydrolyzable analog of allolactose, IPTG, which binds and releases the lac repressor from the lac operator, thereby allowing the initiation of transcription to proceed. It is commonly used in molecular biology as a reporter marker to monitor gene expression.
Repressor proteins bind to the DNA strand and prevent RNA polymerase from being able to attach to the DNA and synthesize mRNA. Inducers bind to repressors, causing them to change shape and preventing them from binding to DNA. Therefore, they allow transcription, and thus gene expression, to take place.
The lacZ fragment, whose synthesis can be induced by IPTG, is capable of intra-allelic complementation with a defective form of β-galactosidase enzyme encoded by host chromosome (mutation lacZDM15 in E. coli JM109, DH5α and XL1-Blue strains). [4] In the presence of IPTG in growth medium, bacteria synthesise both fragments of the enzyme.