Hi all - Illumina is making the MiSeq – acording to their own words – “obsolete”. The MiSeq i100, its replacement, is one of Illumina’s “two-color” (2-channel) instruments. These instruments just use two dyes to identify the 4 bases (A/C/G/T). As a result, no signal from either dye is read as a G, which leads to long stretches of poly-G calls with very high quality scores. My assumption is that minimal changes to the current SOPs are required to analyze i100 data, given mothur’s ability to aggressively filter homopolymers, but I would very much welcome your thoughts.
Hey pad - thanks for your post. I had heard grumblings about this, but have yet to get any data. If anyone sequences a mock community with the i100 I’d love to take a look. But for now, I’d assume that the previous SOP should work well.
Pat
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The other issue I have found with the two color system is intense tag jumping. I would recommend you to do a careful sanity check afterwards before going deep into analysis.
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I see - thank you for the heads-up. So far I have not seen this based on co-sequenced positive and negative controls. How did you reach the tag-jumping conclusion?
I realized that the top SV of each region (we were comparing regions) were also in all other regions. When it would be impossible. Since we were comparing to replicates made in MiSeq, the problem was quite clear, miseq did not have the issue. Important to have negative or mock controls there.
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