We utilize QIIME2 to analyze 16S data. We recently had a pool of bovine nasal swabs sequenced via the new Miseq i100. After importing, quality scores are essentially unavailable. I have attached an image of what that looks like compared to the same sample type on the old MiSeq. Based on another forum post this is likely due to how the new machine categorizes quality scores. Is this issue being addressed? Is the only work around to keep utilizing the old MiSeq? Any guidance is appreciated on how to resolve this issue.
See the below code. We utilized QIIME2 2024.5 via the Advanced Research Computing center at our University.
Yes, these are binned quality scores. Illumina uses this on all their newest machines, maybe because old "unbinned" quality scores were never that accurate anyway?
Good news! The quality scores are there! They are just all just Q37, which is the highest value for these new binned Q scores.
I'm glad you are investigating your new hardware! I've heard lots about Illumina's small machines, so you may soon have your own stories to tell.
Are these plots from the same library sequenced twice, or are these different samples?
How long is your expected PRC product, only 100 bp or so?
EDIT: Here's a GitHub discussion about how these binned q-scores effect error modeling in DADA2. So they can cause issues!
Thank you for the response. I am glad to hear that our sequences are of good quality! Obviously, that is much to my surprise!
These are not the same samples, but this sample type always had lower quality (compared to some of our other sample types) on the old MiSeq. I suppose that’s why I was rather confused at the high values with the i100. We use the V4 region which is about 250bp.
I will look into the resources you sent to familiarize myself with the changes.
