Quantify gDNA in buccal swab - (Nov/13/2012 )
I'm trying to perform PCRs direct on buccal swab material (no lysis, just buccal swab in water then vortex). I fear I'm getting very low DNA yield but what is the quickest way to check DNA quantity? Agarose gel would just be a smear,right?
OD260 would be meaningless with all cell debris RNA etc.?
Sorry if this has been asked before, I've been searching for half an hour and could find anything complete
There isn't really any easy answer to this, your best bet would be to try some sort of fluorescence method, such as the Invitrogen QuantIT system, it might pick up RNA too, but that should typically be less than 5% of the total nucleic acid component of a cell.
DNA binds the cotton swab amazingly tight. Water can only elute ~5% of DNA from a dried specimen swab. We have developed an AquaGenomic method that can fully recover the DNA from dried specimen swabs. You can extract the DNA from the buccal swab with AquaGenomic and then quantify the DNA by conventional methods, from UV spec to real-time PCR.
<*>It yields 5-10 ug of DNA from a dried specimen swab, instead of 200-500 ng.
<*>It improves detection sensitivity.
<*>Its lysate may be used for PCR without further DNA purification.
<*>Dried specimen swabs can be shipped and stored long-term at room temperature.