The GenElute® PCR Clean-Up Kit is designed for rapid purification of single-stranded or double-stranded PCR amplification products (100 bp to 10 kb) from other components in the reaction, such as excess primers, nucleotides, DNA polymerase, oil and salts. This kit combines the advantages of silica binding with a convenient spin column format, eliminating the need for expensive resins or toxic organic compounds such as phenol and chloroform. DNA purification is achieved in a few easy steps. First, DNA is bound to a silica membrane within the spin column. The bound DNA is washed and the clean, concentrated DNA is eluted in the buffer of choice. Each column can purify up to 100 µl or 10 µg of PCR amplified DNA and recover up to 95% of PCR products between 100 bp and 10 kb. More than 99% of the primers and most primer-dimers (<40 bp) and other components are removed. The bound DNA is washed and the clean, concentrated DNA is eluted in the buffer of choice. Purified DNA can be used in enzymatic reactions, including conventional or automated sequencing, cloning and microarray analysis. Purification with the GenElute® PCR Clean-Up Kit takes about 8 minutes. One kit contains reagents sufficient for 70 purifications and offers a convenient storage at room temperature.
We purchased GenElute® PCR Clean-Up Kit to be used as a required step of the whole genome amplification (WGA) procedure. We used the Genomeplex® WGA Kit manufactured by Sigma-Aldrich to perform WGA of old genomic DNA samples stored in our laboratory. The first step of the GenElute® PCR clean-up protocol includes adding Column Preparation Solution to each column followed by centrifugation. Then the total WGA amplification reaction was resuspended in the 5 volumes of the Binding solution, applied to the binding column and centrifuged. After washing with Wash Solution (prepared in advance) and an extra round of centrifugation to remove excess ethanol, samples were collected in 50 ul of the Elution Solution. All solutions were provided in the kit in very generous amounts, volumes far exceeded the minimum required to process 70 samples. Small aliquots of the eluted DNA samples were quantified by measuring absorbance at 260 nm using a spectrophotometer. DNA yields after WGA were very consistent with our expectations. The expected 10 ug of DNA was synthesized from initial 100 ng of genomic DNA. We also ran an aliquot of the amplified and eluted DNA samples on a 1% agarose gel. A bright smear was clearly visible in the 100-1000 bp range, indicating that the procedure worked as expected.
The GenElute® PCR Clean-Up Kit was very easy to work with. The procedure was easy and clearly written in a two-page manual. The only negative point is that during storage at room temperature, a precipitate sometimes forms in the solutions and needs to be dissolved in a warm water bath. It takes five minutes for it to dissolve. Anyone who is doing WGA using one of the several Genomeplex® WGA kits manufactured by Sigma-Aldrich will be required to perform the clean-up procedure with the GenElute® PCR Clean-Up Kit.