Why use Tris saturated phenol instead of directly using phenol?

Release time:

2024-01-04


Introduction

In molecular biology experiments, phenol/chloroform extraction is a commonly used method for DNA extraction. Among them, Tris saturated phenol is a commonly used reagent in this method. However, why not use phenol directly and instead use Tris saturated phenol? This article will explore this issue from multiple perspectives.

Differences in properties between phenol and Tris saturated phenol

1. pH value

Phenol itself is a weakly acidic substance with a pKa value of 9.99. Therefore, without adding any buffer, the pH value of phenol is easily affected by environmental changes, such as changes in pH value during extraction due to contact with acidic or alkaline substances. This change in pH value may affect the efficiency and purity of DNA extraction.

In contrast, Tris saturated phenol is obtained by equilibrating phenol with Tris HCl buffer. During this process, Tris HCl buffer plays a role in stabilizing the pH value, allowing the pH value of Tris saturated phenol to remain stable within a certain range. This stability enables Tris saturated phenol to play a more reliable role in DNA extraction.

2. Solubility

Phenol has a relatively low solubility in water, especially in cold water. This may lead to the problem of phenol crystallization during DNA extraction, thereby affecting extraction efficiency and purity.

In contrast, the Tris component in Tris saturated phenol can increase the solubility of phenol in water, thereby reducing the possibility of crystallization. Meanwhile, Tris components can also form hydrogen bonds with DNA, increasing the stability of DNA in solution and further improving the efficiency and purity of DNA extraction.

Advantages of Tris saturated phenol in DNA extraction

1. Improve DNA extraction efficiency and purity

Due to its stable pH value and high solubility, Tris saturated phenol can more effectively extract DNA from cell lysates. Meanwhile, Tris components can also form complexes with impurities such as proteins, thereby reducing the impact of these impurities on DNA extraction. Therefore, using Tris saturated phenol can improve the efficiency and purity of DNA extraction.

2. Reduce protein pollution

During DNA extraction, proteins are a common pollutant. If ordinary phenol is used for extraction, proteins may be extracted together with DNA, which can affect subsequent experimental results. The use of Tris saturated phenol can effectively reduce protein contamination. This is because Tris components can form complexes with proteins, thereby separating proteins from DNA.

3. Suitable for multiple sample types

Tris saturated phenol is suitable for DNA extraction of various sample types, including bacteria, fungi, animal and plant tissues, etc. This is because Tris components can form hydrogen bonds with DNA from various types of biological samples, enabling effective extraction.

Conclusion

The reason for using Tris saturated phenol instead of directly using phenol is that its stable pH value and high solubility can improve the efficiency and purity of DNA extraction, reduce protein contamination, and be suitable for DNA extraction of various sample types. Therefore, in molecular biology experiments, it is recommended to use Tris saturated phenol for DNA extraction to obtain better experimental results.

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