Nextseq and Miseq data output difference?

Question

[Payal]

Hi,

Is it ok to run a Mixcr pipeline on a TCR sequencing dataset, half of which was processed on Nextseq and half on Miseq? What is the major difference among Nextseq and Miseq platforms?

Thanks, Payal

[GenoMax]

Major difference is the chemistry. NextSeq uses two color where as MiSeq users 4-color chemistry. Hopefully you have data from NextSeq v.2.5 (or what ever the new chemistry is called).

Is it ok to mix sequencing instruments in one analysis .. probably not unless you are able to introduce a batch effect correction (if one exists in software you are using).

You should never change anything in your experimental protocol mid-way during an ongoing experiment.

[h.mon]

Your post lacks information, please add details of the experimental setup. Were all the samples processed uniformly except for sequencing platform? Or were they processed at different dates / library prep kits / laboratories? Are samples distributed randomly among sequencing platform (and other potential batch effects), or were some treatments sequenced exclusively at a single platform? Also, not everybody knows what MiXCR and TCR, so please clarify.

What is the major difference among Nextseq and Miseq platforms?

There are many differences, really. The main one is NextSeq uses the two-dye system, and MiSeq uses the four-dye system.

Answer 1

[Charles Warden]

It doesn't really answer your question about the Mixcr analysis (although I would definitely recommend not just using that strategy for analysis, to maximize your familiarity with the data).

In terms of the general question, maybe it would help to see some information on the Illumina website?

https://www.illumina.com/systems/sequencing-platforms/miseq/specifications.html

https://www.illumina.com/systems/sequencing-platforms/nextseq/specifications.html

https://www.illumina.com/systems/sequencing-platforms.html

For example, the overall throughput is different. So, if you were worried about recombination and/or some amount of sample mixing (emphasizing the value of focusing on the most abundant sequences), maybe that could be indirectly relevant. Or, if you are purchasing your own sequencer, I would guess an iSeq/MiniSeq/MiSeq is more likely than a NextSeq (although some labs can even have their own HiSeqs).

Answer 2

[mizraelson]

From our experience it is better to compare samples sequenced using the same platform. Nevertheless, you can try to use your data derived from both MiSeq and NextSeq. It also depends on the data type, like if your data has UMIs (and new MiXCR version has a UMI support) you can use it to normalize data for more valid comparison.