Similarities and differences between Tris and Tris-HCl and Tris-EDTA

Tris: trimethylol aminomethane

Trismethylaminomethane (Tris) is an organic compound with the formula (HOCH 2) 3CNH 2. Tris is used to prepare buffers in biochemistry and molecular biology experiments. Both TAE and TBE buffers (used to dissolve nucleic acids) used in biochemical experiments require Tris, which can condense with aldehydes when it contains amino groups. Buffering characteristics Tris is a weak base. At room temperature (25°C), its pKa is 8.1. According to buffering theory, the effective buffering range of Tris buffer is between pH 7.0 and 9.2.

The pH of the aqueous solution of Tris base is about 1 0.5. Generally, hydrochloric acid is added to adjust the pH value to the desired value, and then the buffer solution with this pH value can be obtained. But at the same time, the influence of temperature on the pKa of Tris should be controlled.

Since Tris buffer is a weak alkaline solution, DNA will be deprotonated in such a solution, thereby improving its solubility. People often add EDTA to Tris hydrochloric acid buffer to make "TE buffer". TE buffer is used for DNA stabilization and storage. If the pH-adjusted acid solution is replaced with acetic acid, then "TAE buffer" (Tris/Acetate/EDTA) is obtained, and if it is replaced with boric acid, then "TBE buffer" (Tris/Borate/EDTA) is obtained. These two buffers are used in nucleic acid electrophoresis experiments.

1 M Tris-HCl (pH7.4, 7.6, 8.0)

Component concentration 1M Tris-HCl

Preparation volume 1L

Preparation method:

1. Weigh 121.1g Tris into a 1L beaker.

2. Add about 800ml of deionized water and stir to dissolve.

3. Add the amount of concentrated HCl as shown in the table below to adjust the required pH value.

pH Concentrated HCl

7.4 about 70ml

7.6 about 60ml

8.0 about 42ml

4. Bring the solution to 1 L.

5. After high temperature and high pressure sterilization, store at room temperature.

Note: Allow the solution to cool to room temperature before adjusting the pH value, because the pH value of the Tris solution varies greatly with the temperature. When the temperature increases by 1°C, the pH value of the solution decreases by about 0.03 units.

1.5 M Tris-HCl (pH8.8)

Component concentration 1.5M Tris-HCl

Preparation volume 1L

Preparation method

1. Weigh 181.7g Tris in a 1L beaker.

2. Add about 800ml of deionized water and stir to dissolve.

3. Adjust the pH to 8.8 with concentrated HCl.

4. Bring the solution to 1 L.

5. After high temperature and high pressure sterilization, store at room temperature.

Note: The solution should be cooled to room temperature before adjusting the pH value, because the pH value of Tris solution varies greatly with temperature,

For every 1°C increase in temperature, the pH of the solution decreases by approximately 0.03 units.

The advantages of Tris-HCl buffer are:

①Because Tris base is more alkaline, you can use this buffer system to prepare a buffer solution with a wide range of pH values ​​from acidic to alkaline;

② Little interference to biochemical process, no precipitation with calcium, magnesium ions and heavy metal ions.

The disadvantages are:

① The pH value of the buffer solution is greatly affected by the concentration of the solution, the buffer solution is diluted ten times, and the change in pH value is greater than 0.1;

②The temperature effect is large, and the temperature change has a great influence on the pH value of the buffer solution, namely: △pKa/℃=－0.031, for example: the pH of the buffer solution at 4℃=8.4, then the pH value at 37℃= 7.4, so it must be prepared at the use temperature, the Tris-HCl buffer prepared at room temperature cannot be used at 0 ℃ ~ 4 ℃.

③ It is easy to absorb CO2 in the air, so the prepared buffer should be tightly sealed.

④ This buffer solution will interfere with certain pH electrodes, so use an electrode compatible with Tris solution.

Tris solution can absorb carbon dioxide from the air, pay attention to sealing during storage, if you require sterility, you can add sodium azide.

TE is Tris-EDTA buffer (10mM Tris, 1mM EDTA, pH7.4 pH7.6 pH8.0).

Commonly used molecular biology reagents for DNA dissolution.

Prepare 10×TE Buffer (pH7.4, 7.6, 8.0)

Component concentration: 100mM Tris-HCl, 10mM EDTA

Preparation volume: 1L

Preparation method:

1. Measure the following solution and place in a 1L beaker.

1M Tris-HCl Buffer (pH7.4, 7.6, 8.0) 100 ml

500 mM EDTA (pH8.0) 20ml

2. Add about 800ml of deionized water to the beaker and mix evenly.

3. After adjusting the volume to 1L, sterilize at high temperature and high pressure.

4. Store at room temperature.