Agarose Gel Extraction Protocol (QIAEX II)

 

Materials:

Methods:

1. Excise the DNA band from the agarose gel with a clean, sharp scalpel. Minimize the size of the gel slice by removing excess agarose. Collect the gel into a 1.5ml microfuge tube for processing up to 250 mg agarose.
2. Weigh the gel slice in a colorless tube. Add 3 volumes of Buffer QX 1 to 1 volume of gel for DNA fragments 100bp - 4kb.
3. Resuspend QIAEX II by vortexing for 30 sec. Add QIAEX II to the sample according to the table below and mix:

   

4. Incubate at for 10 min to solubilize the agarose and bind the DNA. Mix by vortexing every 2 min to keep QIAEX II in suspension. Check that the color of the mixture is yellow.
5. Centrifuge the sample for 30 sec and carefully remove supernatant with a pipet.
6. Wash the pellet with 500 of Buffer QX1: Resuspend the pellet by vortexing. Centrifuge the sample for 30 sec and remove all traces of supernatant with a pipet. This wash step removes residual agarose contaminants.
7. Wash the pellet twice with 500 of Buffer PE: Resuspend the pellet by vortexing. Centrifuge the sample for 30 sec and carefully remove all traces supernatant with a pipet. These washing steps remove residual salt contaminants.
8. Air-dry the pellet for 10-15 min or until the pellet becomes white. For suspension, air-dry for 30 minutes.
9. To elute DNA, add of 10 mM Tris-Cl, pH 8.5 or and resuspend the pellet by vortexing. For DNA fragments kb, incubate at room temp for 5 min. For DNA fragments 4-10 kb, incubate for 5 min.
10. Centrifuge for 30 sec. Carefully pipet the supernatant into a clean tube. The supernatant now contains the purified DNA.
11. Store the DNA