TOPO TA Cloning (Invitrogen)

Purifying PCR Products, continuedLow-Melt AgaroseMethodPlease note that gel purification will result in a dilution of your PCR product. Use onlychemically competent cells for transformation.1. Electrophorese all of your PCR reaction on a low-melt TAE agarose gel (0.8 to1.2%).2. Visualize the band of interest and excise the band.3. Place the gel slice in a microcentrifuge tube and incubate the tube at 65°C until thegel slice melts.4. Place the tube at 37°C to keep the agarose melted.5. Use 4 µl of the melted agarose containing your PCR product in the TOPO ® Cloningreaction (page 5).6. Incubate the TOPO ® Cloning reaction at 37°C for 5 to 10 minutes. This is to keepthe agarose melted.7. Transform 2 to 4 µl directly into chemically competent One Shot ® cells using themethod on page 6.NotePlease note that the cloning efficiency may decrease with purification of the PCR product.You may wish to optimize your PCR to produce a single band.16