qPCR detection - (Feb/28/2013 )
This might be a stupid question (since I cannot seem to find info about it anywhere), but is there any technical reason not to use SYBR safe instead of SYBR green for qPCR quantification? They have quite similar absorption/emission spectra.
I ask because I want to test some equipment we acquired, and we have plenty of SYBR safe (for gel staining) If I could use this initially instead of buying SYBR green that would be... well to be honest, I could start sooner if I didn't have to wait for SYBR green delivery
Wikipedia (I have no other detailed source of the exact composition of SYBR Safe) says this:
SYBR Green I is another dsDNA stain, produced by Invitrogen. It is more expensive, but 25 times more sensitive, and possibly safer than EtBr, though there is no data addressing its mutagenicity or toxicity in humans.
SYBR Safe is a variant of SYBR Green that has been shown to have low enough levels of mutagenicity and toxicity to be deemed nonhazardous waste under U.S. Federal regulations. It has similar sensitivity levels to EtBr,but, like SYBR Green, is significantly more expensive. In countries where safe disposal of hazardous waste is mandatory, the costs of EtBr disposal can easily outstrip the initial price difference, however.
I got from that, that SYBR Safe is 25 times less sensitive than SYBR Green I used in qPCR, also problems with SYBR are that even after all it is an PCR inhibitor and this issue was surely adressed in it's developement for qPCR. Unlike SYBR Safe, which was obviously designed to be less toxic, which led to decreased sensitivity. Probably if more detailed description of the SYBR Safe molecule was available, we could say more, but I'm affraid it's most likely kind of proprietary formula. So there is unknown inhibitory effect of SYBR Safe plus if you wanted to get the same sensitivity you would need to add more of to the reaction, which would definitelly affect the inhition of PCR.
My opinion: I wouldn't use it for qPCR.
Ok, that makes sense.... Thanks a lot