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Common problems and answers in the preparation of guanidinium isothiocyanate solution

2019-04-10 来源:亚科官网
 
guanidinium isothiocyanate is a powerful protein denaturant which can rapidly dissolve proteins, cause cell structure to break, and nuclear proteins are rapidly separated from nucleic acids due to the disappearance of secondary structure destruction, and have strong denaturing effects on RNase. Thus, guanidinium isothiocyanate can be used in denaturing agents to lyse cells, separate RNA from proteins, and release RNA into solution. However, in the preparation and use of guanidinium isothiocyanate solution, there are often some problems, and this article will answer these questions one by one.
Does the guanidinium isothiocyanate solution need to prepare when used?
The extraction reagent is exemplified by a final concentration of guanidinium isothiocyanate of 4 M: firstly prepared with 42 mM sodium citrate, 0.83% N-lauryl sarcosine (dodecyl group, sodium N-methylglycine), 0.2 mM β-mercaptoethanol to prepare CSB buffer solution; 25 g of guanidinium isothiocyanate, 33 mL of CSB buffer, mixed until completely dissolved, and heated to dissolve at 65 °C to obtain a denaturing solution, which was stored at 4 °C for use.
Therefore, the guanidinium isothiocyanate solution may not need to be prepare when used, and there are also different concentrations of the solution on the market, but the placement time is not too long, otherwise there will be precipitation, and need to be carefully checked before use; If too much is prepared at one time, it is recommended to do the subpackage.
Is it necessary to sterilize the guanidinium isothiocyanate solution after it is formulated?
The guanidinium isothiocyanate solution is not necessarily sterilized, the solution is not very stable, and the lysate is generally treated with 0.1% DEPC water, and the water has been sterilized. In addition, all reagents for RNA extraction are formulated with DEPC water, and the reagents are preferably newly opened and used exclusively for RNA.
Although the prepared lysate does not need to be sterilized, it should be noted that the plastic products used in the experiment have been marked Rnase-Free, if they have not been used, they can be used directly. Others must be processed: DEPC was added to the double distilled water to a final concentration of 0.1%. The plastic product was immersed therein, and overnight in fuming cupboard, then sealed with aluminum foil, autoclaved for at least 30 minutes, and dried for use.
What is the toxicity of guanidinium isothiocyanate?
Inhalation, ingestion, skin contact with guanidinium isothiocyanate can cause damage, so wear lab coat, gloves and goggles when handling; contact with acid can release extremely toxic gases, harmful to aquatic organisms, it possiblely has long-term adverse effects on the water environment and should therefore be released into the environment.
How to deal with the used waste liquid?
The pollution of the laboratory mainly includes biological pollution and chemical pollution, which can be roughly classified into wastewater, exhaust gas and solid pollutants according to the form. Among them, biological pollution includes biological waste pollution and biological bacterial toxin pollution, while chemical pollution includes organic pollution and inorganic pollution. In general, all kinds of waste liquids need to be classified. In principle, the original bottles are recycled. If mixing is required, it is necessary to determine that the reagents do not generate heat, toxic gases, explosions, etc. after mixing. The name and concentration of the drug should also be indicated on the recycling bottle for uniform recycling, and should not be discarded, resulting in environmental pollution and poisoning.
With the rapid development of molecular biology, genetic diagnosis has become the most accurate and effective means of detection. In the process of genetic diagnosis, the extraction of viral nucleic acid is a prerequisite for detection, and the quality of nucleic acid extraction also affects the accuracy of detection. The quality of the lysate directly affects the quality of the nucleic acid extraction, which in turn affects the results of the experiment. As the most commonly used cell lysate, guanidinium isothiocyanate solution can only obtain the best experimental results based on safety experiments by processing every detail.
Related links: guanidinium isothiocyanate
Edited by Suzhou Yacoo Science Co., Ltd.