Internal controls/ Reference Samples for Nanopore

Question

[pixie@bioinfo]

Hello, we have an in-house Minion and mostly work with plant viruses and viroids. Since flowcells vary in their sequencing capacity, is there a way to use internal controls/reference samples to compare across runs ? Would like to know advantages and disadvantages of the same. Thanks

[GenoMax]

Trying to imagine how these will work. You can't be sure that the controls will always behave in reliable/predicted way since they will be subject to same uncertainty as the rest of the sample for a given flowcell.

[dthorbur]

I agree with GenoMax, and I'm unsure if this would actually be informative since there are more factors at play than just capacity. Quality and throughput also vary dynamically across runs as pores get clogged or stop functioning and this can be due to the quality of the input and things like contamination.

Feasibly, you could add multiple spike-ins as a control, but what would you do with that information if you do see variation among runs? It seems unnecessary in most use cases to remove sequences from one run because another had lower capacity if you went the normalization route.