How much RNA is needed for labelling and how much labelled cDNA is needed for hy - (Aug/28/2006 )
I would like to know how much RNA is needed for labelling and how much labelled cDNA is needed for hybridization, thanks a lot!!
For Affymetrix, usually starting with 5-6 ug total RNA (high quality, check 200 ng on 1.2% agarose gel. You have to see two and only the two expected bands and nothing below).
For hybridisation usually 15 ug adjusted (minus input cDNA) cRNA is used.
With this values it works well, may be it is possible to use less, never tried.
all depends of your organism, and your labelling technique.
For example, with bacteria you could label up to 20 µg of RNA with a direct or an indirect ( without amplification ) labelling.
But if you are working with biopsy or little sample ( ARNT 3µg) , you need to use an amplified labelling method ( a lot of kit exist).
If you sample is very limited, try Illumina's BeadArray which only needs 500ng total RNA as starting material.
just like the post above, which platform? even for affymetix, there are a few labeling options and how much total rna can you extract.