The ChargeSwitch® Direct gDNA Kit is a solid phase, high-throughput system for genomic DNA (gDNA) extraction. Under low ionic conditions, DNA binds to the sides of the ChargeSwitch® plate, allowing for proteins, salts, and other contaminants to be washed away. Each well allows for the binding of ~50 ng of gDNA which facilitates the performance of downstream reactions directly in the plate. Alternatively, you can elute your DNA for use in multiple downstream reactions.
The kit comes with the ChargeSwitch® plate and four different buffers. The process is very simple and suitable for automated liquid handling platforms. Simply mix 10 ìl of cell lysate, whole blood, or cell-free sample with 100 ìl of Binding and Lysis Buffer and incubate at room temperature for 30 minutes. Aspirate the liquid, wash the wells twice with Wash Buffers, and you’re done! Either add your PCR master mix directly to the wells, or elute the DNA with 100 ìl of Elution Buffer for later use.
Before choosing this kit, I evaluated several different DNA isolation kits including silica-based methods using vacuum or spin columns, ultrafiltration techniques, and magnetic bead technology. I based my decision on cost, ease of use, likelihood of cross contamination, throughput, and DNA yield. This kit prevailed by a large margin. The cost is significantly lower than most of the other methods. Also, I could perform the entire procedure in one biological safety cabinet, using a 96-channel liquid handler, an 8 channel liquid dispensing system, and a plate mixer. Many of the other techniques that I tested required centrifugation, pouring off a filtrate, or transferring samples to several different plates, which can increase the risk for cross-contamination or sample mix-up. The entire ChargeSwitch® process is performed in one plate, and microsatellite profile comparisons between input DNA and purified DNA products using ProfilerPlus® showed no evidence of contamination or cross-contamination between wells. My throughput was about six 96-well plates every two hours, and yields were comparable to the other systems.
I used this kit for the extraction of DNA from ~7,300 crude cell lysates. Although the kit is designed for the user to perform the downstream application directly in the plate, I chose the optional elution method. This allowed me to perform several reactions with one extract. I was at first concerned about the 10-fold dilution of the DNA during elution (my final concentrations were between 0.5 and 5 ng/ìl), but I found that the extracted DNA was of such high quality that I could obtain reliable results on sensitive assays using just 1 ìl of the elution. The product was successfully used to perform Taqman®, STR analysis, and even a long-range duplex PCR assay.
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Jacqueline Huckins
ORISE Fellow
Molecular Biology Branch, DLS, NCEH
Centers for Disease Control and Prevention
United States
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