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Ligation Transformation problem ???

Are you purifying your bands (nicely visible on the gel) and ligating/transforming, but disappointed that next day 'NO COLONY' has grown on your plates?? Well, one very common source of the problem resides in the way you purify your bands. If you run your bands in a gel with Ethidium Bromide (Eth. Br.) then cut your band under UV light, that should be your problem. Follow these tips: - Use a gel without Eth. Br. - Load your product1 in as many consecutif (neighboring) wells as needed - Load around 50-100 ng (just enough to be able to see it under UV, after Eth. Br. staining) of product1 right next to a DNA size marker - Do the same if you have more than one product - Run your gel as usually, and when finished, cut the gel portion that has the Marker and the one well with the minimum product1 - Stain with Eth. Br., and rinse - Visualize under UV and take a picture (to make sure the bands are there at the correct size) - With a scalp mark the top and bottom of your band (s) of interest - Take this stained portion of the gel and place it back next to the unstained gel - Following the marks you made, cut the band (that you don't see) from the unstained gel - purify using your preferred gel purification Kit - Ligate, transform, now you should get your colonnies