PCR Purification

For PCR purification, we used the E.Z.N.A.™ Cycle-Pure Kit. This kit is used to purify PCR products, as well DNA products in enzymatic reactions, particularly, restriction digests. We followed the manufacturer’s Spin Protocol.

Reagents and Materials

• E.Z.N.A.™ Cycle-Pure Kit
• PCR products

Protocol

1. Determine the volume of the PCR reaction, transfer the sample into a clean 1.5ml microcentrifuge tube, and add 4-5 volumes of Buffer CP
2. Vortex thoroughly to mix
3. Spin down tubes shortly
4. Place a HiBind DNA column in a provided 2ml collection tube
5. Add 100µl Equilibration Buffer to the column
6. Incubate at room temperature for 4 minutes
7. Spin at maximum speed for 20 seconds
8. Apply the sample to the HiBind DNA column and centrifuge at 10,000x g for 1 min at room temperature
9. Discard liquid
10. Wash the HiBind DNA column by adding 700µl of DNA Wash Buffer diluted with absolute ethanol and centrifuge centrifuge at 10,000x g for 1 min at room temperature
11. Discard liquid and repeat last step with 500µl of DNA Wash Buffer
12. Discard liquid and centrifuge the empty HiBind DNA column for 2 min at maximum speed to dry the column matrix
13. Transfer HiBind DNA column to a clean 1.5mL tube and elute with 30-50µL of ddH2O
14. Use a spectrophotometer to measure the concentration and the purity of your plasmid