RNA library preps

Prep. type	Min. amount	Recommend amount	Max. volume	Accepted buffers
mRNA   total RNA	0.5 μg total RNA or purified mRNA / rRNA-depleted RNA / bacterial RNA	1 - 10 μg total RNA or 10 - 400 ng mRNA purified from 1- 10 μg starting total RNA	50 μl   5 μl per 1 μg starting material	TE, 10 mM Tris, H20   H20

Input requirements for RNA library preps.

We offer TruSeq^TM stranded RNA-seq sample prep as a service, which employs poly-T beads to enrich the polyadenylated fraction of mRNA. If the use of poly-T beads is not appropriate for your sample, it is your responsibility to inform us of this. Alternative kits/procedures also exist for rRNA depletion, but these are not offered as a standard service by the NSC. We may consider performing these as part of a research collaboration – please inquire. 

For bacterial RNA / rRNA depleted eukaryotic RNA: You may perform rRNA depletion yourself and submit the rRNA-depleted or mRNA-enriched samples to us to enter the RNA-seq prep at the appropriate stage in the protocol. It is critical that the RNA is in this case supplied only in water.  We can also perform RNA-seq on total RNA without RNA depletion in this way, although you must be prepared that the majority of reads will be derived from rRNA.

Note that we cannot guarantee successful prep of rRNA-depleted samples, as it is usually impossible to distinguish successfully depleted samples from degraded RNA. These library preps are therefore performed at the user's risk.

We strongly recommend that RNA be prepared by procedures that include a DNase digestion to remove contaminating DNA.