Nucleic acid purification for use with PCR, quantitative real time PCR (qPCR) and the isothermal DNA amplification methods loop-mediated DNA amplification (LAMP) and recombinase polymerase amplification (RPA).
Recommended Usage
Pipette 100 μL of Extraction Buffer into a 1.5 mL tube. Add 1-2 mm3 of a sample to the tube.
Grind the sample with a small plastic pestle, and dilute the sample with an additional 400 μL of Extraction Buffer. Dip a dipstick up and down 3 times into the extract, then dip 5 times into 1 mL of wash buffer. Discard the wash buffer. Keep the dipstick to release the DNA in the following steps.
Dip the dipstick into 20–50 µl PCR reaction mix 3–15 times (~10 s total) to release the DNA.
Alternatively to store the DNA sample, dip the dipstick into a tube of TE buffer. If necessary, molecular grade water can also be used for storage.
Limitations
The dipstick purification method does not significantly concentrate the DNA sample, unlike solid-phase nucleic acid techniques. This means the method is best suited to PCR-based applications, and not suitable for methods such as restriction enzyme digests or clean up of PCR amplicons.
Due to the small capture volume of the dipstick, this method is unsuitable for purifying large quantities of nucleic acids, which means it is not suited to methods such as genome sequencing that require larger quantities of DNA For multiple PCR reactions from the same sample, an individual dipstick will be required.
Components
Extraction Buffer (20 mM Tris-HCl, 25 mM NaCl, 2.5 mM EDTA, 0.05% SDS, 2% PVP-40, pH 8)
Wash Buffer (10 mM Tris-HCl, pH 8)
DNA Dipsticks (filterpaper, wax)
Storage & Stability
Store at room temperature (18–25 °C) for up to 1 year. For longer term storage, store at 4 ºC or freeze at –20 ºC.
Shipping conditions
Shipped at room temperature.
