Dual rnaseq normalization

Question

[slapiiv]

Hi everybody, I have performed a dual rnaseq (infected cells by a pathogen) at different points, at the beginning host-corresponding reads are more than 90% of the total, however, at a later point, I have almost 50% or reads corresponding to the host and 50% corresponding to the protozoo (total reads are the same on average). I have divided the NGS data into two files (host and protozoo) for analyzing expression with DEseq2. One step of deseq2 is precisely the read depth coverage (RDC) normalization but in this specific case, I do not know if this normalization is enough for dual rnaseq where I have these huge RDC differences. Any advice or special recommendation?

Answer 1

[grant.hovhannisyan]

If you have successfully separated the reads of the host and pathogen, and have the count table for each species, then you can proceed with a standard DESeq2 analysis. If you have time-series design, you should follow the instruction in DESeq2 vignette https://www.bioconductor.org/packages/devel/bioc/vignettes/DESeq2/inst/doc/DESeq2.html#time-series-experiments

For reference https://link.springer.com/protocol/10.1007%2F978-1-4939-7634-8_4

[slapiiv]

Nice, I will take a look but it seems very useful, thanks.