Isopropylthio-β-galactoside (IPTG) is a molecular analogue of isopentose that is widely used in molecular biology. Isopentose is a lactose metabolite that triggers transcription of the lac manipulator. Therefore, it can be used to induce genes controlled by the lac operon to express proteins, such as E. coli proteins. IPTG induction, on the other hand, is a method to regulate protein synthesis by triggering the transcription of the lac operon. IPTG induction in bacteria typically involves both fast and slow induction, with fast induction not being applicable to all proteins and may result in suboptimal yields. Slow induction can improve the solubility of some proteins, depending on the protein you are studying and the application. Most importantly, IPTG concentration also affects the induction results, so you need to screen for the right induction concentration.

Induction concentration

The commonly used IPTG induction concentration in the laboratory ranges from 100 μM to 1.0 mM, and under this condition, protein expression can be effectively induced. Higher concentrations of IPTG may be required if mutant lacIq, which overproduces lac repressors, is present.

After integrating the Pgrac100 promoter into the lacA locus of the B. subtilis genome, the results of the integrated inducible expression system showed that the Pgrac100 promoter was able to tightly control the expression of the target proteins, with the induction factors after 4 h induced by 0.01, 0.1, and 1 mM isopropyl β-D-1-thiogalactopyranoside (IPTG) being 3.3, 7.6, and 10.8. The length of induction time and the concentration of IPTG had a significant effect on the expression effect of the target protein. Specifically, the expression of GFP increased significantly after 4 h of induction with 0.1 mM IPTG, accounting for 13% of the total cellular protein.

How to make a IPTG – 1 M (100 x) Stock Solution

(1) Weigh 2.383 g of IPTG.

(2) Add 10 ml sterile H2O. Dissolve completely.

(3) Prewet a 0.2 μm syringe filter by drawing through 5-10 ml of sterile H2O and discard water.

(4) Sterilize Ampicillin Stock through the prepared 0.2 μm syringe filter.

Dispense the prepared stock solution into 1 ml portions and store at -20°C for up to 1 year. Dilute to the required concentration for each use and dispense as required.

References:

[1] CLARA L KIELKOPF  Ina L U  William Bauer. Expression of Cloned Genes in E. coli Using IPTG-Inducible Promoters.[J]. Cold Spring Harbor protocols, 2021. DOI:10.1101/pdb.prot102137.

[2] CHU T B P, NGUYEN D, PHAN T P T. Development of expression vector with the Pgrac100 promoter induced by IPTG and integrated gfp+ gene into Bacillus subtilisgenome at lacA locus[J]. Ministry of Science and Technology, Vietnam, 2024. DOI:10.31276/vjst.66(1).75-80.

[3] SEAVER S. IPTG – 1 M (100 x) Stock Solution[C]. 2014. DOI:10.17504/protocols.io.ci3ugm.