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Old 02-27-2019, 04:34 AM   #1
Adiro
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Location: Israel

Join Date: Jan 2019
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Default ERCC spike-ins for prokaryotes

Hi!
I see that many people use ERCC spike-ins with prokaryotes, but looking at the manufacturer description I see that they only mention that it is good for eukaryotes... https://www.thermofisher.com/order/c...roduct/4456740

I found some articles that asses the validity of the ERCC in eukaryotes, but never on prokaryotes...

So can I use it in prokaryotes? anyone found an article assessing the validity of ERCC in prokaryotes?


**I'm specifically interested in e.coli, and I really need spike ins because I suspect that most of the genes will express differently between the two conditions... If for example all the genes will be 2 times more expressed in condition A than in condition B, without spike-ins the differential expression will look like there was no change. If you have other suggestions for this scenario besides ERCC, I would love to know... **
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Old 02-27-2019, 05:33 AM   #2
Bukowski
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I don't think they say it's only good for eukaryotes, just that it mimics the nature of eukaryotic transcripts in terms of nucleotide content, size distribution, polyadenylation etc. I think the bigger concern using them for prokaryotes is probably that a chunk of the ERCC spike ins are derived from bacterial sequences in the first place.

If you're concerned about the derivation of the ERCC sequences, then there are artificial spike-in's available: https://www.rna-seqblog.com/spliced-...g-experiments/ (I would link to the sequins site, but there appears to be a configuration issue on their webserver)

Given that you "already see many people use ERCC spike-ins with prokaryotes" what kind of additional validity are you looking for?
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Old 02-28-2019, 04:51 PM   #3
cmbetts
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I'd be careful about using them for Prokaryotes. Many of them are actually bacterial genes, mostly B.subtilis if I remember correctly, but also some antibiotic resistance genes descended from Affy controls. I'd download the fastas and align to your genome of interest before using them.
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