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Forum: Illumina/Solexa 08-01-2017, 07:19 AM
Replies: 113
Views: 18,526
Posted By GW_OK
There've been a few hypotheses that ExAmp is...

There've been a few hypotheses that ExAmp is actually Recombinase Polymerase Amplification (RPA), developed by TwistDX.

Here's a Youtube video describing it (https://youtu.be/x6AbsOrSAoY?t=1m17s)...
Forum: Illumina/Solexa 03-28-2017, 06:55 AM
Replies: 113
Views: 18,526
Posted By GW_OK
I don't know if you can truly compare...

I don't know if you can truly compare efficiencies of the ExAmp chemistry with the other instruments.

On the HiSeq and NextSeq instruments you are randomly clustering across the flowcell with a...
Forum: Illumina/Solexa 03-20-2017, 01:39 PM
Replies: 113
Views: 18,526
Posted By GW_OK
A few more interesting points. The tile map...

A few more interesting points.

The tile map for the S2 flowcell:
-The first digit is the lane number: 1 or 2.
-The second digit represents the surface: 1 for top or 2 for bottom.
-The third...
Forum: Illumina/Solexa 03-20-2017, 08:39 AM
Replies: 113
Views: 18,526
Posted By GW_OK
Maybe we need to start talking in terms of...

Maybe we need to start talking in terms of "library coverage" instead of "genome coverage"...

Also, don't confuse read numbers with cluster numbers. There'll be 2 reads to every cluster. And on...
Forum: Illumina/Solexa 03-20-2017, 07:41 AM
Replies: 113
Views: 18,526
Posted By GW_OK
Regarding sampling depth, I am dubious. The...

Regarding sampling depth, I am dubious.

The Truseq PCR-free protocol (which I have to assume they're using) has you start with 1ug for 350bp inserts and 2ug for 550 bp inserts. Since they say...
Forum: Illumina/Solexa 03-20-2017, 06:52 AM
Replies: 113
Views: 18,526
Posted By GW_OK
It's from the BaseSpace project NovaSeq: WGS...

It's from the BaseSpace project NovaSeq: WGS TruSeq PCR-Free 450 (6plex) So I reckon it must be PCR-free.

NA12878-rep1 only looking at data from lane 1.
Forum: Illumina/Solexa 03-20-2017, 06:36 AM
Replies: 113
Views: 18,526
Posted By GW_OK
So, yeah. I didn't miss that qcfail blog post....

So, yeah. I didn't miss that qcfail blog post. I've read through it several times. I wanted to recapitulate their analysis on a data set that:
(A) had not undergone PCR amplification and
(B) was...
Forum: Illumina/Solexa 03-20-2017, 06:25 AM
Replies: 113
Views: 18,526
Posted By GW_OK
Finally, I graphed duplicate read coordinates...

Finally, I graphed duplicate read coordinates relative to the initial read coordinates (x1-x2/y1-y2 from the file I made above). I've attached that file below since it's not tremendously large. Most...
Forum: Illumina/Solexa 03-20-2017, 06:17 AM
Replies: 113
Views: 18,526
Posted By GW_OK
I then ran clumpify markduplicates=t...

I then ran clumpify markduplicates=t dupedist=2500 spantiles=t to coalesce the duplicates I used a simple perl script to parse the fastq headers into a "tile1 x1 y1 tile2 x2 y2" text...
Forum: Illumina/Solexa 03-20-2017, 06:12 AM
Replies: 113
Views: 18,526
Posted By GW_OK
The forums aren't letting me post a big post so...

The forums aren't letting me post a big post so I'm going to break this into three posts.

I've been intrigued with the question of duplicate-well directionality. Does it follow the direction of...
Forum: Illumina/Solexa 03-18-2017, 07:20 AM
Replies: 113
Views: 18,526
Posted By GW_OK
deleted due to duplication (hah)

deleted due to duplication (hah)
Forum: Illumina/Solexa 03-16-2017, 05:46 PM
Replies: 113
Views: 18,526
Posted By GW_OK
Well, since I have a 3000 (but no extant PCR-free...

Well, since I have a 3000 (but no extant PCR-free data) I wanted to look at the public 4000 data from Basespace, specifically:
NA12878-PCRfree450_S3_L003_R1_001.fastq.gz...
Forum: Illumina/Solexa 03-16-2017, 07:06 AM
Replies: 113
Views: 18,526
Posted By GW_OK
Can anyone share exactly how they're getting X/Y...

Can anyone share exactly how they're getting X/Y coordinates from the patterned flowcell fastq? I'm only seeing a single number, which I am guessing corresponds to a well ID.
Forum: Illumina/Solexa 03-14-2017, 06:37 AM
Replies: 2
Views: 761
Posted By GW_OK
2xxx tiles are on the bottom surface. I concur...

2xxx tiles are on the bottom surface. I concur with the possibilities raised by Markiyan. Looks like it started somewhere around cycle 35. Definitely a call to tech support is in order.
Forum: Illumina/Solexa 02-07-2017, 06:55 AM
Replies: 6
Views: 2,297
Posted By GW_OK
Yep. Works fine.

Yep. Works fine.
Forum: Illumina/Solexa 01-10-2017, 11:50 AM
Replies: 113
Views: 18,526
Posted By GW_OK
Eco tweeted it. And I think he was there at the...

Eco tweeted it. And I think he was there at the presentation?

Anyway, I can't find confirmation either way on the official Illumina pages.
Forum: Illumina/Solexa 10-20-2016, 08:41 AM
Replies: 13
Views: 3,761
Posted By GW_OK
One thing we have found to be helpful is to clean...

One thing we have found to be helpful is to clean up and run the qPCR products out on our Tapestation. This way you know exactly what size of product you have in your analysis.
Forum: Illumina/Solexa 07-26-2016, 08:00 AM
Replies: 4
Views: 1,050
Posted By GW_OK
Also if you do any bisulfite work or anything...

Also if you do any bisulfite work or anything with low base diversity the 4000 is not nearly as robust as the 2500.
Forum: Illumina/Solexa 05-12-2016, 07:02 AM
Replies: 7
Views: 615
Posted By GW_OK
In the same way that the machine knows that each...

In the same way that the machine knows that each cluster of light forms the bases for each read: the pixels encompassing each light signal are precisely mapped. When the machine "finds" the clusters,...
Forum: Illumina/Solexa 03-16-2016, 08:29 AM
Replies: 6
Views: 1,496
Posted By GW_OK
Those would be the Mid kits, which intrinsically...

Those would be the Mid kits, which intrinsically have lower throughput.

Going back and looking at the data more closely, the "collapse" in Q30 seems to be mostly the Mid kits.
Forum: Illumina/Solexa 03-16-2016, 07:26 AM
Replies: 6
Views: 1,496
Posted By GW_OK
We shoot for loading at 1.6pM. It seems to work...

We shoot for loading at 1.6pM. It seems to work very well for us. We've gone up to 1.8, and down to 1.4, but we usually go back to 1.6.

Here are some quick graphs from 40 past runs showing how...
Forum: Illumina/Solexa 02-25-2016, 11:10 AM
Replies: 8
Views: 1,002
Posted By GW_OK
Sounds about right, then. I wouldn't run...

Sounds about right, then.

I wouldn't run 2x300's on the Miseq, though, if you're planning on doing that.
Forum: Illumina/Solexa 02-25-2016, 11:01 AM
Replies: 8
Views: 1,002
Posted By GW_OK
I don't know the context of Illumina's statement,...

I don't know the context of Illumina's statement, so I can't comment.
Forum: Illumina/Solexa 02-25-2016, 10:44 AM
Replies: 8
Views: 1,002
Posted By GW_OK
An Illumina sequencer will, or at least should,...

An Illumina sequencer will, or at least should, generate the same amount of reads for every run, respective to the chemistry loaded on it. A v3 Miseq kit should give about ~20M reads per run.
...
Forum: Illumina/Solexa 01-20-2016, 06:44 AM
Replies: 33
Views: 4,625
Posted By GW_OK
I completely agree. I am puzzled, though,...

I completely agree.

I am puzzled, though, why any Core would not immediately have had the run replaced by Illumina and reloaded if it did not meet spec.
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