RNA sequencing methods are available to match your project goals for gene expression, detection of alternative gene expression, splicing, and budget.
3’ mRNA sequencing
|Reads per sample (millions)||2–5||30||60|
|Hands-on time (hours)||1.75||1.5||3.5|
|Total time (hours)||4.5||4.5||6.5|
|Input range||100 pg–500 ng*||1–25 ng**||100–1000 ng|
|Species compatibility||3' mRNA Blood: Human|
3' mRNA: All
* Inputs of >200 ng are recommended for efficient detection of low abundance transcripts.
** rRNA depleted or mRNA enriched RNA
Total RNA-Seq offers the most comprehensive whole transcriptome analysis. mRNA-Seq is ideal if the research is focused only on the coding region and limited amounts of starting material are available.
Variation in RNA-seq expression data can be attributed to a variety of factors ranging from the quality of the starting material to the person performing the experiment. The External RNA Controls Consortium (ERCC), an ad-hoc group of academic, private, and public organizations hosted by the National Institute of Standards and Technology (NIST), developed a common set of external RNA controls to distinguish between these sources of variability and true gene expression. The controls consist of a set of unlabeled, polyadenylated transcripts designed to be added to an RNA sequencing experiment after sample isolation in order to measure against defined performance criteria.
The Invitrogen ERCC Spike-In Mix is a pre-formulated blend of 92 transcripts, derived and traceable from NIST-certified DNA plasmids. The transcripts mimic natural eukaryotic mRNAs of 250 to 2,000 nt in length. Inclusion of ERCC controls in RNA sequencing experiments establishes a standard measure for data comparison across gene expression experiments and makes it possible to measure sensitivity (lower limit of detection) and dynamic range of an experiment.
For Research Use Only. Not for use in diagnostic procedures.