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Troubleshooting: Protein Expression
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IPTG
Product Description
IPTG is an analog of galactose that is non-metabolizable and inactivates the lac repressor to induce synthesis of β-galactosidase in E. coli. The expression of cloned genes under the control of the lac operon is induced by IPTG. It is also a substrate for thigalactoside transacetylase and has been reported to induce penicillinase in bacteria.
IPTG is commonly used in cloning procedures that require induction of β-galactosidase and is most often used with X-Gal (Gold Bio #X4281C) or Bluo-Gal (Gold Bio #B-673) for blue/white colony screening or Magenta-Gal (Gold Bio #B-378) for red/white colony screening of bacterial colonies.
IPTG is also used in the induction of recombinant proteins. In those systems, a protein of interest is encoded downstream of the IPTG inducible promoter. In the presence of IPTG, the protein of interest is induced in the cell culture. The culture can then be lysed and the protein expressed and purified through a number of methods, including His Tag or GST purification systems (for proteins with ligand tags).
GoldBio IPTG is made from a synthetic, non-animal origin.
Product Specifications:
IPTG
Isopropyl-beta-D-thiogalactoside
MOLECULAR BIOLOGY GRADE
Forumla: C9H18O5S
MW: 238.30 g/mol
Purity: >99% by HPLC
Dioxane free
Storage/Handling: Store desiccated at -20°C. Protect from light.
PubChem Chemical ID: 656894
IPTG Quick Answers:
How does IPTG induce the lac operon?
IPTG, which is similar to the lactose metabolite allolactose, induces the lac operon by binding to the lac repressor. Once IPTG binds to the lac repressor, a conformational change occurs, causing the lac repressor to dissociate from the lac promoter. Once the lac repressor dissociates, RNA polymerase binds to the promoter to initiate expression.
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How much IPTG should be used for expression/ What concentration of IPTG should be used for expression?
Most papers recommend using between 0.1mM – 1.0 mM IPTG for expression. However, the amount you should use depends on your proteins. However, it is best to run an IPTG titration with ranges between 0.01mM – 1.0mM and compare using SDS-PAGE to determine the optimal concentration. Something to keep in mind is higher concentrations of IPTG can cause inclusion bodies.
How do you dissolve IPTG?
IPTG can be dissolved in sterile water.
To make fresh 100mM IPTG stock solution:
- Weigh 0.238 g of IPTG
- Add 10 ml sterile H2O. Dissolve completely.
- Prewet a 0.22 µm syringe filter by drawing through 5-10 ml of sterile H2O and discard water.
- Sterilize IPTG Stock through the prepared 0.22 µm syringe filter.
- Store in 1 ml aliquots at -20°C for up to 1 year.
How do you store IPTG?
Store powder IPTG desiccated at -20°C. Protect from light. For IPTG stock solution, store in 1 ml aliquots at -20°C for up to 1 year.
What is IPTG used for?
Primary applications of IPTG include:
- Inducing proteins
- Blue-white screening using X-Gal
IPTG Tutorial Videos:
Product Specifications
| Catalog ID | I2481 |
|---|---|
| Name(s) |
IPTG Isopropyl-beta-D-thiogalactoside |
| CAS # | 367-93-1 |
| Formula | C9H18O5S |
| MW | 238.30 g/mol |
| Grade | MOLECULAR BIOLOGY GRADE |
| Storage/Handling | Store desiccated at -20°C. Protect from light. |
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