DNA Extraction from Agarose Gel

QIAquick Gel Extraction, a Simplified version of the Protocol

Step 1: Cut out your DNA band

Use a clean scalpel to carefully cut the desired DNA fragment from the agarose gel.

Step 2: Weigh the gel slice

Place the gel piece into a clean microcentrifuge tube and weigh it accurately.

👉 For every 100 mg of gel, add 300 µl of Buffer QG (3 volumes).

Note

This means for every 100 mg of gel, add 300 µl Buffer QG.

Example:
• 100 mg gel → add 300 µl Buffer QG
• 200 mg gel → add 600 µl Buffer QG
• 300 mg gel → add 900 µl Buffer QG

Step 3: Dissolve the gel

Heat the tube at 50°C for 10 minutes, vortexing every few minutes until the gel is completely melted and the solution turns yellow.

If it looks orange/purple instead, add 10 µl sodium acetate (3M, pH 5.0) and mix until yellow.

Note

Sodium acetate adjusts the pH so that DNA binds properly to the column during purification.

Step 4: Add isopropanol

Add 1 gel volume (for example, about 100 µl for 100 mg gel) of isopropanol and mix thoroughly.

Note

Isopropanol increases DNA binding efficiency to the silica membrane in the QIAquick column.

Step 5: Bind DNA to the column

Insert a QIAquick spin column into the provided 2 ml collection tube.

Load your sample onto the column and spin for 1 minute.

If your sample is larger than 800 µl, load it in two rounds using the same column.

Discard the flow-through after each spin.

Step 6: Remove impurities

Add 500 µl Buffer QG, spin for 1 minute, and discard the flow-through.

Add 750 µl Buffer PE, spin for 1 minute, and discard the flow-through again.

Spin once more (with an empty tube) for 1 minute to dry the column completely.

Step 7: Elute DNA

Place the column in a clean 1.5 ml microcentrifuge tube.

Add 50 µl Buffer EB (10 mM Tris-Cl, pH 8.5), pre-warmed to 55°C, directly onto the membrane.

Let it stand for 1–4 minutes, then spin for 1 minute to collect your purified DNA.

Note

Pre-warming Buffer EB helps increase DNA recovery from the membrane.

(Optional) Check DNA on a gel

If you want to verify your DNA, mix 1 part loading dye with 5 parts purified DNA and load it on an agarose gel.