What final library concentration do you consider "good" for an Illumina RNA TruSeq library? 10nM?
If you presume the 15 cycles of amplification recommended by the kit produces 1.8x increase of amplicons per cycle, then I think your initial number of amplicons (unamped library molecules) could be as low as 9 million. At least that is what I get when I run the calculations. (Presuming the final amped libraries is eluted in 10ul.)
That may result in an acceptably low amount of PCR duplicate in a 10 million read data set, but not so great for 30 million.
Anyone else surprised by this?
(Please check my math, if you are so inclined.)
Comments?
--
Phillip
If you presume the 15 cycles of amplification recommended by the kit produces 1.8x increase of amplicons per cycle, then I think your initial number of amplicons (unamped library molecules) could be as low as 9 million. At least that is what I get when I run the calculations. (Presuming the final amped libraries is eluted in 10ul.)
That may result in an acceptably low amount of PCR duplicate in a 10 million read data set, but not so great for 30 million.
Anyone else surprised by this?
(Please check my math, if you are so inclined.)
Comments?
--
Phillip
Comment