So there may not be many of us left, but those who are still using 454-Junior machines... have you had as many problems with the upgrade to long reads as we have had? Before the upgrade, we never had a failure, now we can barely get a "+" run to pass specs. Sometimes the library looks whacky (technical term for a few different results). Sometimes the library looks okay but the run has very few reads that pass the filters.
The vast majority of the problem shows up as very high Trimmed too short, and this is followed by high Mix+Dots/Total Key pass. We've been getting advice from Roche, but mostly they are blaming the emPCR conditions. We've varied the conditions and even gone back to the old Titanium chemistry and run settings without much success. We don't have any FLX machine (or experience) so we aren't mixing those protocols that I know of. I tried a few configuration hacks for the filters, but nothing worked.
I'd like to hear of a couple success stories so we can emulate what you have done...
The vast majority of the problem shows up as very high Trimmed too short, and this is followed by high Mix+Dots/Total Key pass. We've been getting advice from Roche, but mostly they are blaming the emPCR conditions. We've varied the conditions and even gone back to the old Titanium chemistry and run settings without much success. We don't have any FLX machine (or experience) so we aren't mixing those protocols that I know of. I tried a few configuration hacks for the filters, but nothing worked.
I'd like to hear of a couple success stories so we can emulate what you have done...
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