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Forum: Illumina/Solexa 06-29-2017, 01:20 AM
Replies: 2
Views: 742
Posted By matth431
Thanks - it seems to have generated a makefile OK...

Thanks - it seems to have generated a makefile OK just running bcl2fastq with --use-bases-mask set to y101nnnnnn (since it had processed up to base 107), will know for sure once the make script has...
Forum: Illumina/Solexa 06-28-2017, 05:54 AM
Replies: 2
Views: 742
Posted By matth431
Data recovery from aborted HiSeq2500 run

Hi, we were running a 1x126bp high-output run on our HiSeq, but a comms issue caused the run to fail at cycle 109. Basecalling was complete however for cycles 1-107, although for some reason Q30%...
Forum: Illumina/Solexa 01-14-2016, 03:04 AM
Replies: 26
Views: 15,661
Posted By matth431
MadsAlbertsen, would you mind reposting a link to...

MadsAlbertsen, would you mind reposting a link to the hack protocol? Your previous link has now removed the hack details as "the extra reagent in the v3 kit makes it unnecessary"... however recently...
Forum: Illumina/Solexa 01-20-2015, 01:01 AM
Replies: 53
Views: 72,383
Posted By matth431
I asked Illumina and was told not. Apparently the...

I asked Illumina and was told not. Apparently the cameras are more sensitive.

Also, the NextSeq550 capability for arrays is currently limited to the methylation and karyomapping chips, not sure...
Forum: Illumina/Solexa 11-26-2014, 04:55 AM
Replies: 3
Views: 1,946
Posted By matth431
What version of MCS is the MiSeq running? We had...

What version of MCS is the MiSeq running? We had issues with recent runs until Illumina suggested upgrading to v2.5.

We do still see issues with some samples - one user experiences a massive...
Forum: Sample Prep / Library Generation 11-26-2014, 04:52 AM
Replies: 2
Views: 2,547
Posted By matth431
It's ultrapure water, so also DNase and RNase...

It's ultrapure water, so also DNase and RNase free if you want to save on buying Sigma Water (since we always have a massive excess of Illumina PW1 left over).

But yeah, PW just stands for Pure...
Forum: Illumina/Solexa 11-26-2014, 04:49 AM
Replies: 1
Views: 1,617
Posted By matth431
The rapid cluster kit provides all of the...

The rapid cluster kit provides all of the reagents needed to cluster the library onboard the HiSeq2500 machine - the diluted library goes into the FCA or FCB template position to the left of the...
Forum: Illumina/Solexa 11-26-2014, 04:39 AM
Replies: 6
Views: 3,073
Posted By matth431
Yes, would be keen to know this. I'm also...

Yes, would be keen to know this. I'm also disappointed Illumina haven't taken the opportunity to introduce 300-cycle kits as we have a lot of users who want 2x151 to help with polyploid read mappings...
Forum: General 11-25-2014, 02:13 AM
Replies: 0
Views: 1,409
Posted By matth431
BluePippin issue

Hi,

After a few successful runs on our BluePippin, it is now repeatedly failing calibration. There's nothing in the manual about what to do if calibration fails - SageScience's support pages are...
Forum: Illumina/Solexa 11-25-2014, 02:10 AM
Replies: 10
Views: 2,279
Posted By matth431
One reason we've done this is where there's a...

One reason we've done this is where there's a high proportion of short fragments in the library - we've had a user make Nextera libraries from very low amounts of viral DNA which tend to...
Forum: Illumina/Solexa 05-26-2014, 01:36 AM
Replies: 1
Views: 1,525
Posted By matth431
Lifespan of TruSeq SBS reagents once defrosted

We routinely store incoming SBS kits in the freezer (they're shipped on dry ice) and I've been advised previously that the reagents are only good for ~15 days once thawed. I've never actually noticed...
Forum: Illumina/Solexa 08-02-2013, 05:35 AM
Replies: 4
Views: 5,269
Posted By matth431
We won't be doing this routinely, and a set of...

We won't be doing this routinely, and a set of fusion primers with 50+ barcodes would set us back ~5000.
Forum: Illumina/Solexa 07-25-2013, 04:26 AM
Replies: 4
Views: 5,269
Posted By matth431
Thanks, that's interesting. We do end repair and...

Thanks, that's interesting. We do end repair and A-tailing on size-selected libraries purified via the LabChip XT system, so based our input for this amplicon library on the recovery from that, which...
Forum: Illumina/Solexa 07-25-2013, 01:37 AM
Replies: 4
Views: 5,269
Posted By matth431
PCR-free libraries

Hi,

I think I've figured out a problem we have with a recent experiment. We have amplicons of ~400bp with a 5bp index added on to both the 5' and 3' ends (part of the PCR primers). The forward and...
Forum: Illumina/Solexa 05-08-2013, 03:54 AM
Replies: 17
Views: 9,270
Posted By matth431
Our kits ran out in August 2012 and are still...

Our kits ran out in August 2012 and are still working fine, just click "Proceed anyway" when challenged on the RFID.
Forum: Illumina/Solexa 04-30-2013, 07:05 AM
Replies: 2
Views: 1,566
Posted By matth431
Coming back to this for anyone who observed a...

Coming back to this for anyone who observed a similar problem - we had lane blockages so fluorescence was building up. Eventually, the sequencer gave up scanning those lanes but luckily it was close...
Forum: Illumina/Solexa 04-22-2013, 05:15 AM
Replies: 2
Views: 2,225
Posted By matth431
Ours gives very good data when operational - most...

Ours gives very good data when operational - most of our downtime has been caused by fluidics issues as (unlike HiSeq) these are an addition to the machine (the SQ module) rather than being an...
Forum: Ion Torrent 04-19-2013, 11:44 PM
Replies: 9
Views: 5,648
Posted By matth431
This was a bit delayed as the Torrent Server...

This was a bit delayed as the Torrent Server doesn't come pre-loaded with the hg19 reference sequence, but our last control run was similar in output to the one I mentioned above and was aligned...
Forum: Ion Torrent 03-28-2013, 04:54 AM
Replies: 9
Views: 5,648
Posted By matth431
We've tried a couple of the test libraries now...

We've tried a couple of the test libraries now and seem to be making progress. Last run gave an average of 89bp read length with 180bp being the maximum. We got 42M reads (about 3.7Gbp). Our issue...
Forum: Illumina/Solexa 03-18-2013, 06:40 AM
Replies: 20
Views: 20,978
Posted By matth431
I wasn't aware it was possible to restart a...

I wasn't aware it was possible to restart a failed MiSeq run? Surely it rejects the cartridge and flowcell based on the RFID tags?
Forum: Illumina/Solexa 03-18-2013, 03:23 AM
Replies: 20
Views: 20,978
Posted By matth431
I just had a similar "Object reference" failure...

I just had a similar "Object reference" failure about 185bp into read 1. In our case, cluster density was very low (Nextera XT normalised libraries pooled and diluted as per instructions but...
Forum: Illumina/Solexa 03-05-2013, 03:33 AM
Replies: 37
Views: 20,333
Posted By matth431
Interesting that folk are seeing a difference pre...

Interesting that folk are seeing a difference pre and post-upgrade. We've just run our first v2 kit (though not first v1) since upgrade at the usual 8pM and obtained broadly the same density we...
Forum: Illumina/Solexa 03-05-2013, 03:31 AM
Replies: 23
Views: 8,574
Posted By matth431
Just got 10.3Gbp raw data, histogram suggested...

Just got 10.3Gbp raw data, histogram suggested 7.0Gbp >Q30 by end of 251|6|251 run. Useable sequence (post-QC trim) checking it on Genomics Workbench probably closer to 9Gbp. About 39.5M reads with...
Forum: Illumina/Solexa 02-15-2013, 05:00 AM
Replies: 18
Views: 13,559
Posted By matth431
That's good to know - unfortunately, we can't do...

That's good to know - unfortunately, we can't do rapid chemistry as yet due to only having a HiScanSQ. We are looking at upgrading in the next few months to a 2500, but the user is a bit more...
Forum: Illumina/Solexa 02-14-2013, 04:00 AM
Replies: 18
Views: 13,559
Posted By matth431
Hi DocBio, Could I check whether you ever...

Hi DocBio,

Could I check whether you ever perform mixed flow-cells with some lanes TruSeq and some Nextera (not both in same lane)? We carried out a run last year with six lanes of TruSeq and 2...
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