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Old 05-04-2011, 06:26 AM   #1
pmiguel
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Default Sequencing Analysis Viewer trick.

Illumina's "Sequencing Analysis Viewer" is a useful windows client for monitoring the progress of a run. Interestingly if a single sequence dominates in a lane against a background of mixed sequences, you can actually read that dominant sequence from an SAV window. The one that seems to work the best is on the "Analysis" tab, in the "Data By Cycle" window. If you choose "% Base" from the top drop down menu and the lane of interest, you might see something like:



This is from an ongoing training run. We constructed some TruSeq DNA libraries and wanted to at least try our hands with some RNA seq. We did not have the TruSeq RNA kit on hand, so we just took some cDNA we had made using a SMART kit for an earlier 454 run.

If you read the sequence from the plot above it matches that of a 454-modified SMART adapter. (See the "interrupted oligo-T"--even the terminal "V" is evident? Well we are reading the other strand.) Clearly would need some work on the library construction methodology not to waste so many bases on adapter sequence. But for a training run...

We spiked the cDNAs into an indexed run lane that was about 1/2 non-cDNA to avoid focus issues.

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Old 05-04-2011, 08:20 AM   #2
kmcarr
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Quote:
Originally Posted by pmiguel View Post
Illumina's "Sequencing Analysis Viewer" is a useful windows client for monitoring the progress of a run.
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Phillip
Looking at the post run IVC plots it was always easy to pick out the samples containing even modest amounts of primer dimer this way. Now I can see them during the run; still can't do diddley squat about other than to tell the lab folks.
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