Why are the SIRV isoforms available as 3 mixes?

In SIRV-Set 1, each mix of E0, E1, and E2 contains all 69 SIRV isoforms in different concentration ratios.

• The concentration ratios in Mix E0 are identical (1:1). This Mix is ideal for assessing the detection capabilities of a given RNA-Seq workflow, since all 69 transcripts are present in equimolar concentration, thus leading to an unbiased detection regardless of sequencing depth. 

• Mix E1 covers one order of magnitude (up to 1:8), and E2 extends over more than two orders of magnitude (up to 1:128). This allows the user to either assess all SIRV transcripts spiked-in at the same level or to mimic the transcript variant concentration distribution encountered in real samples.

Moreover, the comparison of 3 samples, each spiked with either E0, E1, and E2 allows for a detailed assessment of differential gene expression on the transcript level.

The inter-mix concentration ratios range from 1/64- to 16-fold.

The final concentration of the SIRV Isoform Mixes contain identical mass concentrations of 25.2, 25.2, and 25.4 ng/µl, and molar concentration of 69.0, 68.5 and 70.8 fmol/µl for mixes E0, E1 and E2 respectively.