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Application of IPTG (CAS 367-93-1) and detection in biochemical samples

2018-11-09 来源:亚科官网
In recent years, molecular diagnostics has become a popular branch in the field of IVD, and its detection targets are nucleic acids and proteins, mainly nucleic acid molecules. In the biochemical samples that need to be prepared, IPTG is often present. IPTG is expensive, and when it is used too much, it is not only toxic, but also increases production costs. Therefore, the detection of IPTG content in products is very important. This article will summarize the application of IPTG and introduce a new method for detecting IPTG.
IPTG is an activity-inducing substance of β-galactosidase. It has the following effects:
      (1) Selection of recombinants: When the vector DNA of the pUC series is transformed with lacZ-deficient cells as the host, or when transfected with the vector DNA of M13 phage, if X-gal and IPTG are added to the plate medium, genetic recombination could be selected based on whether white colonies (or plaques) are present because of the α-complementarity of β-galactosidase;
      (2) Use as an expression inducer of an expression vector having a promoter such as lac or tac. As a molecular biological reagent, IPTG is commonly used for blue-white spot screening and IPTG-induced protein expression in bacteria. In genetic engineering technology, IPTG is used as an inducer to promote the expression of a large amount of protein of interest.
How to detect its content? IPTG is actually a semi-thio compound. Due to the lack of luminescent groups in the molecules of sulfur-containing compounds, it cannot be detected by conventional UV-Vis detectors. The most commonly used detection method is derivatization combined with fluorescence HPLC. Detection. However, this method is time consuming and labor intensive and has poor selectivity. Therefore, the development of an IPTG content detection method to achieve accurate determination of IPTG content in biochemical samples has positive practical significance. Researchers have developed a method for detecting IPTG content, including the following steps:
(1) Pretreatment: ultrafiltration centrifugation (the ultrafiltration tube has a pore size ranging from 0.001 to 0.02 μm), and a sample to be tested having a molecular weight of less than 3000 Daltons is obtained;
(2) IPTG standard curve drawing: standard solution was prepared and detected by ion chromatography-pulse amperometric detection. The column is a reversed phase column; the mobile phase is a mixed mobile phase of a buffer solution (pH 1 to 11) and an organic solvent (volume ratio is 99 to 80: 1 to 20), and an IPTG standard curve is obtained;
(3) Detecting the sample to be tested: ion chromatography-pulse amperometric detection method, and comparing the standard curve, the specific content of IPTG is obtained.
The method uses an ultrafiltration centrifuge tube to process the sample, and quickly and efficiently remove impurities in the system, and the treated sample solution is separated into a reverse phase chromatography column through a mixed mobile phase of a pH buffer solution and an organic solvent, and the IPTG is quickly washed. Take it out and test it by pulse amperometric detector to complete the analysis. The sample is easy to pre-treat, has good specificity and high sensitivity, and is suitable for popularization and application.
Reference
[1] Wang Li. A method for detecting IPTG content. 2017, CN106353445A.
Related links: IPTG
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Edited by Suzhou Yacoo Science Co., Ltd.