I wish to normalize a scRNASeq dataset with respect to ERCC spike-ins, where "for some of the samples, ERCC spike-in RNA was added to the lysis buffer" and was wondering how to do so?
I have seen similar questions posted on the forum, such as the following: Normalization methods with RNA-Seq ERCC spike in? and have checked out the package RUVSeq in the top answer. However, in this package, it seems that spike-ins are taken as transcripts, whereas in the dataset I'm analyzing it seems as though 'spike-ins' are respective of the cell being sequenced rather than a transcript. In addition, in my research I stumbled upon the following tutorial for preprocessing of scRNASeq data: https://f1000research.com/articles/5-2122/v2 where also, as in the RUVSeq package, "ERCC" is
grep'd through the
rownames to identify ERCC transcripts.
Which leads me to my second question: what is a spike-in if it is not a transcript - and therefore not within the
rownames of the dataset and rather within the samples? Please note that I am not from a biological background and, I have read the wikipedia article, along with many others, in my quest to understand what a spike-in is.
In addition, please note that I have a metadata file that states exactly which of the samples have these spike-ins and which dont.
Any advice on how to proceed would be much appreciated.