Search Product
Structure Search
Search
Online Support
-
Customer service
Advantage Products
Location: Technique Support
Instructions of IPTG
2012-08-03
来源：亚科官网
Name: isopropyl-β-D-thiogalactoside (IPTG)
Applicationss: IPTG is a highly effective inducer, which is not metabolized by bacteria and very stable, it is widely used in laboratories. IPTG is commonly used in cloning experiments that require the induction of β-galactosidase activity. It is often used in combination with X-Gal or Bluo-Gal for blue-white screening of recombinant bacterial colonies that can induce the expression of the lac operon in E. coli. IPTG binds to the lacI repressor protein and alters its conformation, preventing the inhibition of the lacZ gene encoding β-galactosidase.
How to use: First, IPTG is formulated into an aqueous solution of 23.83 mg/mL (100mM) and filtration, sterilization. Then 100μL of the above solution, 200μL of X-Gal (20mg/mL in dimethylformamide (DMF) solution) and 100 μL of Amp (100mg/ml) were added to 100mL of agar medium, Made into IPTG, X-Gal, Amp plate medium. After the DNA fragment has been inserted into the pUC series vector (or other vector carrying lacZ, Amp gene) and then transformed into lacZ-deficient cells, the IPTG, X-Gal and Amp plate media described above are applied. According to the blue-white, and easily selected gene recombinants (white for DNA recombinants with DNA inserts).
Preparation: Dissolve 2g IPTG in 8mL distilled water, dilute to 10mL with distilled water, sterilize and filter with 0.22μm filter, divide into 1mL aliquot and store in -20℃.
Advantage:
1. Gratuitous inducer, which can induce the synthesis of enzymes, but not be decomposed. Although lactose can induce enzyme synthesis, it also can be decomposed, resulting in a lot of complex kinetic problems, so researchers often use gratuitous inducer to carry out various experiments.
2. IPTG is not metabolized by bacterial cells. It is lactose analogue. Once introduced into cells, IPTG induces long-lasting effects. Therefore, the induction rate of IPTG is very high. Only a very small amount (1 mmol/L) of IPTG is needed to achieve the ideal inducing effect. Because IPTG is not metabolized, it specifically induces the expression of foreign proteins in the cell, not affected by cell metabolism, induction effect is continued stability. Lactose has a double effect, both can be as an inducer of lactose precursor substances, but also can be used as a carbon source in the induction process. IPTG is suitable for experimental study because of its excellent stability.
3. IPTG can be effectively copied without the lacY gene.
4. Sulfur in Galactoside bonds take the place of oxygen, and lost the hydrolytic activity, but the thiogalactoside and homologous oxygen compounds has the same affinity with the enzyme site, although IPTG is not identified by β-galactosidase, but it is a very effective inducer of the lac gene cluster.
Note: When culturing phage, add 25 μL/3mL(24mg/mL) to Top agar. Medium contained IPTG should be stored at 4℃ and dark place, and it should be used within 1 to 2 weeks.
Storage: Store at 2~8°C. Keep away from heat and oxidants.
