Can I pre-amplify a gene of interest using TaqMan gene expression assay for that particular gene from Thermo? If so, do I have to use their TaqMan pre-amp master mix, which is $900 or can I use any other master mixes that our lab already uses (but never been used on TaqMan gene expression assay, which is primer+probe set). Our lab normally uses Q5 hot start high-fidelity master mix from New England biolabs, but we only pre-amplified using primers only rxns, not primer + probe set like TaqMan assays.
Is this for a one off assay? if so you might as well save time, take a bit of a hit on the cost of reagents and just use the Taqman mastermix.
If not.......Do you know the primer sequences for the taqman assay? if so then you can use any polymerase/PCR mix to preamplify. If you don't know the sequence you're going to have spend a bit of time generating cDNA and using various overlapping primer sets along the gene (unless you can be absolutely sure of generating long cDNA transcripts.....how long is the gene?) to see if you can amplify the region (probably toward the 3' end) on which the Taqman primer/probe set sits. Slightly labourious I know but if this is for a long term assay then it'll be worth it
yes, you can use TaqMan gene expression assay to pre-amplify the gene of interest. For the master mix, I would suggest using the Taqman Master Mix. By doing this, you will save a lot of time that otherwise would be spent on optimization etc.
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