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Forum: Illumina/Solexa 01-24-2018, 11:15 PM
Replies: 23
Views: 4,697
Posted By HeinKey
perhaps this question is asked before, but I...

perhaps this question is asked before, but I don't understand the added value of dual indexing over single indexing anymore if both indices are unique in dual indexing. The highest possible...
Forum: Illumina/Solexa 04-06-2017, 05:32 AM
Replies: 1
Views: 751
Posted By HeinKey
Hi anglabat1, I think it is save to randomly...

Hi anglabat1,

I think it is save to randomly remove reads from the over sequenced samples. They are randomly devided over the flow cell, and ordered on pixel position. So taking the same first...
Forum: Sample Prep / Library Generation 11-02-2016, 01:01 AM
Replies: 14
Views: 8,841
Posted By HeinKey
If restriction fragmented DNA can not be sized...

If restriction fragmented DNA can not be sized using Ampure beads, then sheared DNA can also not be sized using these beads.
Since this is a standard procedure I don't think a 4 nt single stranded...
Forum: Illumina/Solexa 07-13-2016, 03:42 AM
Replies: 5
Views: 1,998
Posted By HeinKey
We have the same issue. Low quality index2 in...

We have the same issue. Low quality index2 in paired-end runs, and normal quality index2 in Single read runs.
Illumina finally recognized the problem, (we did some validation runs for them).
We...
Forum: Illumina/Solexa 04-29-2015, 06:20 AM
Replies: 22
Views: 2,409
Posted By HeinKey
We store the library tubes at -20oC. Storing...

We store the library tubes at -20oC.
Storing all "raw" data consequently is far more expensive than occasionally redo a HiSeq lane.
We have re-run 2 years old libraries without any problem.
Forum: Illumina/Solexa 04-29-2015, 06:02 AM
Replies: 7
Views: 2,551
Posted By HeinKey
hi Runsheng, I understood you compared an 800...

hi Runsheng,
I understood you compared an 800 nt library qPCR result with a 500 nt library, and corrected loading with 5/8.
This seems logical given both libraries cluster equally efficient.
This...
Forum: Illumina/Solexa 02-24-2015, 06:38 AM
Replies: 13
Views: 3,620
Posted By HeinKey
we had no problems at all with the delivery of...

we had no problems at all with the delivery of ProClin300 by Sigma.....Is the request for this chemical too high since the introduction of V4 chemistry??
Forum: Illumina/Solexa 01-22-2015, 05:48 AM
Replies: 53
Views: 58,423
Posted By HeinKey
just visited an Illumina meeting in Brussels. ...

just visited an Illumina meeting in Brussels.

Current HiSeqs are not upgradable to the 3000 or 4000 versions. Hardware is too different between the systems.

4000 and 3000 can only sequence...
Forum: Illumina/Solexa 01-15-2015, 04:20 AM
Replies: 53
Views: 58,423
Posted By HeinKey
Can these new flow cells still be overloaded, or...

Can these new flow cells still be overloaded, or has Illumina created the ideal clustering procedure? Just load your library at high concentration for maximal occupance of the flow cell "wells" and...
Forum: Illumina/Solexa 01-13-2015, 04:56 AM
Replies: 53
Views: 58,423
Posted By HeinKey
#SNPsaurus: I assume it will be 2x 150 bp in...

#SNPsaurus:
I assume it will be 2x 150 bp in 3.5 days.
2.1 to 2.5 billion clusters per flow cell, in a 2x150 bp run is 630 - 750 Gb.
With a speed >200Gb per day a 2x 150 bp run will take 3.5...
Forum: Illumina/Solexa 12-04-2014, 05:45 AM
Replies: 9
Views: 1,644
Posted By HeinKey
We have been running low diversity libraries from...

We have been running low diversity libraries from the early GA times on. We used a dedicated lane with Phi-X to calculate the metrix values. This is no longer necessary with the newest RTA versions...
Forum: Illumina/Solexa 12-11-2013, 05:56 AM
Replies: 20
Views: 14,340
Posted By HeinKey
Hi Phillip, For MiSeq runs I agree, but is RTA...

Hi Phillip,
For MiSeq runs I agree, but is RTA for HiSeq also changed? I was told the improvement for biased libraries was only for MiSeq RTA.

Hein
Forum: Illumina/Solexa 11-13-2013, 06:17 AM
Replies: 4
Views: 2,075
Posted By HeinKey
Illumina mentioned you can sequence PE libraries...

Illumina mentioned you can sequence PE libraries on a PE flow cell, in SE mode. So sequencing only read one.
SE dual indexing is possible, also on a PE flow cell. This is because the second index...
Forum: Illumina/Solexa 10-08-2013, 05:28 AM
Replies: 2
Views: 1,910
Posted By HeinKey
27 bases might be too long. The "stuffer" between...

27 bases might be too long. The "stuffer" between P5 and flow cell surface is probably shorter.
I would give it a try on a MiSeq nano run which is fairly cheap to test it. If it works it should be...
Forum: Illumina/Solexa 05-29-2013, 02:52 AM
Replies: 8
Views: 7,637
Posted By HeinKey
Hi Austinso, If you use low complexity pooling...

Hi Austinso,
If you use low complexity pooling strategies be aware to have sufficient balance in each index cycle.
Choose the indices properly, making sure the combination has 50% A/C and 50% G/T...
Forum: Illumina/Solexa 05-29-2013, 02:05 AM
Replies: 3
Views: 2,031
Posted By HeinKey
MikeNice, If you flush your system with high...

MikeNice,

If you flush your system with high salt to remove the air, and then continue, and repeat the second index?
You first need to tweak the HCS so it will image the dark cycli resulting in...
Forum: Illumina/Solexa 04-29-2013, 07:34 AM
Replies: 11
Views: 7,361
Posted By HeinKey
primer hybridization?

Hello Agent99,
If I understand correctly the issue of poor read3 is due to the index (read2) still present on your strands.
I can't understand how this would happen since reclustering takes place...
Forum: Illumina/Solexa 04-25-2013, 05:24 AM
Replies: 4
Views: 2,149
Posted By HeinKey
Hi Anguy, Thnx for your response. I agree...

Hi Anguy,
Thnx for your response.
I agree with you that local disturbances would result in cluster density variation. I checked images and cluster density plots. This all looks normal with equally...
Forum: Illumina/Solexa 04-25-2013, 04:58 AM
Replies: 4
Views: 2,149
Posted By HeinKey
IDDan; thx for your response. We see cluster...

IDDan; thx for your response.
We see cluster density variation within a single flow cell. Variation between lanes loaded from the same diluted single stranded library. Mixed well!
If NaOH is...
Forum: Illumina/Solexa 04-16-2013, 03:25 AM
Replies: 4
Views: 2,149
Posted By HeinKey
Question cluster density variation

Since a few weeks we occasionally observe very big variation between raw cluster densities on our Single read flow cells.
Even if we load multiple lanes from the same library tube we sometimes see...
Forum: Illumina/Solexa 04-16-2013, 03:03 AM
Replies: 4
Views: 1,914
Posted By HeinKey
We are sequencing libraries with mixed fragment...

We are sequencing libraries with mixed fragment lengths for years now. Works fine in our hands. Images are looking more messy, but read quality is ok. Our fragments vary between 200-700 nts in single...
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