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Old 01-13-2011, 06:58 PM   #1
ashchin
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Default Has Anyone stopped a HiSeq run due to low intensity clusters?

I am sorry ,

I meant to ask if anyone has stopped the Hiseq run because of low intensity of clusters using normal stop for keeping it in the safe mode.
I did know how to post a new thread so posted here.
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Old 01-14-2011, 05:14 AM   #2
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Not for that reason, but we have used normal stop to interrupt one run so that we can load a second flow cell. It does not seem to adversely affect the data quality.
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Old 01-14-2011, 08:52 AM   #3
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Yes. If the intensities in the first base report are low, or after 4/5 cycle, the values are still low we have stopped the run. Try re-hybridizing the sequencing primer. If this is across the board (and re-hybridizing doesn't help), let your FSE know. Our lasers were out of alignment.
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Old 01-17-2011, 02:27 PM   #4
ashchin
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Thank you all
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Old 01-17-2011, 07:23 PM   #5
HESmith
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See this link for recommendations about library quantification.
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Old 01-20-2011, 12:30 PM   #6
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Hi,

Have you guys ever stopped the run after the first read?
Or do you know if it is okay to store the flow cell after the first read?

Thank you,
Ashwini
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Old 01-20-2011, 12:33 PM   #7
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It is fine to store the flowcell after read 1 if you want to stop the run. However, if you do want to sequence it eventually, you should re-hybridize the sequencing primer. Just make sure you pump High Salt Buffer through the flowcell before removing it and store it in the fridge. You can store the flowcell this way for at least a month.
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Old 01-20-2011, 12:48 PM   #8
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thank you,
what if I want to run read 2?
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Old 01-20-2011, 12:56 PM   #9
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What do you mean by run read2? If you store the flowcell after first base report you will need to rehybridize the primer and start from cycle 1 and go through the run as normal. I don't think you can store after read 2 in the same way. You can pump buffer through and let sit on the machine for up to 2 days and then reprimer hyb the primers. However, if you remove the flowcell from the machine, you loose ability to relate the read 1 clusters to the positions of read 2.
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Old 01-20-2011, 01:13 PM   #10
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Thank you, I got your point. You mean the clusters might change position when we remove it after read 1.
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Old 01-25-2011, 04:03 PM   #11
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what if read 2 is not giving signal, can I re-prime?
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