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Old 09-19-2012, 12:10 PM   #1
brineybs
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Default Frankenstein MiSeq 2x400bp runs

Has anyone tried running beyond the 2x250bp "official" capacity of the MiSeq? We worked with a sequencing collaborator who hacked together a 2x250 run before the reagents were available by unofficially (although aided by Illumina tech support) pooling a 2x150 and a 1x100 cartridge. Any idea if pooling of a 2x250 and a 2x150 is possible, allowing early access to the supposedly upcoming 2x400 run?
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Old 09-19-2012, 06:34 PM   #2
snetmcom
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I'm not sure you would want to try that yet.

see posts below.
http://seqanswers.com/forums/showthr...t=20455&page=2
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Old 09-20-2012, 05:05 AM   #3
GenoMax
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2 x 250 runs are not stably working (in our hands) as yet. Perhaps others have had better luck. No problems getting sequence, it is the decline in quality over the later cycles.

Hopefully this is just teething troubles that will get sorted out over time.

If you have the time (and money) to try the 2 x 400, go right ahead. Let us know what happens.
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Old 09-20-2012, 05:38 AM   #4
pmiguel
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Quote:
Originally Posted by GenoMax View Post
2 x 250 runs are not stably working (in our hands) as yet. Perhaps others have had better luck. No problems getting sequence, it is the decline in quality over the later cycles.
Is this with a hardware-upgraded instrument? Does going to lower loading densities appear to help?

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Old 09-20-2012, 06:21 AM   #5
TonyBrooks
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http://pathogenomics.bham.ac.uk/blog...-day-2-tweets/

"Illumina are testing 400bp single-end reads in R&D on the MiSeq, with the longest achieved overlapping paired-read of 678bp"

I think I read somewhere that it was around 80% >Q20 @ 400bp
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Old 09-20-2012, 06:35 AM   #6
GenoMax
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Quote:
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Is this with a hardware-upgraded instrument?
Yes. Our instrument has received the hardware upgrade.

Quote:
Does going to lower loading densities appear to help?
--
Phillip
Not sure. v.1. runs (2 x 25 bp) are looking fine, it is the v.2. runs (2 x 250 bp) that seem to be the problem. We have done two long runs so far (one sample and one phiX).

We are working with illumina at the moment. This may be a specific issue with our instrument. We shall see.
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