SEQanswers

Go Back   SEQanswers > Search Forums


Showing results 1 to 25 of 61
Search took 0.02 seconds.
Search: Posts Made By: hoytpr
Forum: Illumina/Solexa 05-15-2019, 05:35 AM
Replies: 0
Views: 633
Posted By hoytpr
Chloroplasts sequenced on NextSeq?

We have a client who wants to sequence chloroplasts (plastids) but from my reading, it seems like he's going to need longer reads than the NextSeq can produce. Am I correct?

The TL;DR part.
He...
Forum: Illumina/Solexa 03-29-2019, 07:44 AM
Replies: 4
Views: 571
Posted By hoytpr
Thanks. Our analysis is off-line also, so we're...

Thanks. Our analysis is off-line also, so we're re-processing now. Appreciate the reply!
Peter
Forum: Illumina/Solexa 03-29-2019, 06:37 AM
Replies: 4
Views: 571
Posted By hoytpr
Thanks for the reply. These are four runs...

Thanks for the reply. These are four runs completed a couple weeks ago, and there are bcl files (e.g. 0001.bcl.ngzf) in four directories
\Data\Intensities\BaseCalls\L001 (through L004). Each .bcl...
Forum: Illumina/Solexa 03-29-2019, 05:51 AM
Replies: 4
Views: 571
Posted By hoytpr
How to get raw reads from NextSeq

We have used bcl2fastq on our local server for years, but a client has asked for untrimmed unfiltered raw reads. What's the easiest way to get these from a completed run, and still get demultiplexed...
Forum: Illumina/Solexa 01-08-2019, 09:29 AM
Replies: 5
Views: 827
Posted By hoytpr
Must just be primer dimers. I talked with...

Must just be primer dimers. I talked with Illumina today and they looked at it. Basically said it's normal for a NextSeq to have these runs of "N" (they call "N-masking when multiplexing") when...
Forum: Illumina/Solexa 01-07-2019, 12:03 PM
Replies: 5
Views: 827
Posted By hoytpr
The % NoCall looks a little weird: ...

The % NoCall looks a little weird:

https://www.dropbox.com/s/osm8higjbe93460/No-Calls.png?dl=0
Forum: Illumina/Solexa 01-07-2019, 11:54 AM
Replies: 5
Views: 827
Posted By hoytpr
Most everything looked okay. There is a little...

Most everything looked okay. There is a little glitch at 35bp in the data by Cycle Error rate

https://www.dropbox.com/s/ijb1ikhr9xahv1t/metrics.png?dl=0
Forum: Illumina/Solexa 01-07-2019, 09:26 AM
Replies: 5
Views: 827
Posted By hoytpr
High % "N" in first 35bp using MultiQC

Hi all,
Just did some sequencing for a client and got a little odd plot using MultiQC for the "FastQ per base N-Content". It looks like the first 35 bp had problems with "N" then suddenly stopped....
Forum: Core Facilities 11-29-2018, 10:16 AM
Replies: 1
Views: 3,201
Posted By hoytpr
Ramiro, you're gonna need a query-able database....

Ramiro, you're gonna need a query-able database. If MySql is not on your favorites list, Microsoft Access has a business template that can be hacked to run a core facility. It's a pain to transfer...
Forum: Illumina/Solexa 11-29-2018, 10:07 AM
Replies: 2
Views: 1,533
Posted By hoytpr
Hi cmccabe, I know this is a late reply, but cat9...

Hi cmccabe, I know this is a late reply, but cat9 is pretty much how any Nextseq500 is hooked up if you aren't using BaseSpace. Our server was set up with an Windows VM and them mapped as a drive to...
Forum: Illumina/Solexa 11-20-2018, 07:32 AM
Replies: 1
Views: 734
Posted By hoytpr
Rolling circle dsDNA vs ssDNA experience?

Does anyone have experience with samples using rolling circle amplification that can tell me whether they have relatively high or low amounts of ssDNA? The client is not local so it would help to...
Forum: Sample Prep / Library Generation 09-13-2018, 06:05 AM
Replies: 3
Views: 1,733
Posted By hoytpr
We just did a Zoom call with them, and I was very...

We just did a Zoom call with them, and I was very impressed. We will order one of their kits when we get in a large number of bacterial DNAs. They are working on RNA kits, but don't offer them yet....
Forum: Illumina/Solexa 08-27-2018, 08:37 AM
Replies: 6
Views: 1,634
Posted By hoytpr
Right, I suspect the final data might be...

Right, I suspect the final data might be affected, but in this case, the project was probably at least a year old. We just want to help.

We have both a Tapestation and 2100 and they tend to agree...
Forum: Illumina/Solexa 08-27-2018, 05:55 AM
Replies: 6
Views: 1,634
Posted By hoytpr
UCan'tBcereus: Your post could not have been...

UCan'tBcereus: Your post could not have been better timed. We have a client with some very precious samples that show both peaks, but the RIN scores are very low. We were considering turning down the...
Forum: Illumina/Solexa 07-25-2018, 06:28 AM
Replies: 6
Views: 1,634
Posted By hoytpr
Thanks for the great advice Jakob. Pete

Thanks for the great advice Jakob.

Pete
Forum: Illumina/Solexa 07-03-2018, 12:54 PM
Replies: 6
Views: 1,634
Posted By hoytpr
Opinion needed on Truseq vs Scriptseq

So quite simply, when doing the math, 48 bacterial RNA-seq samples (libraries) would cost over a thousand bucks more using Truseq + Ribozero, versus using ScriptSeq "complete" + Scriptseq "barcodes"...
Forum: Illumina/Solexa 02-13-2018, 07:53 AM
Replies: 5
Views: 1,168
Posted By hoytpr
So, you'd have to quantify by QPCR then, as...

So, you'd have to quantify by QPCR then, as fluorescence or Bioanalyzers won't tell you the number of correctly formatted sequences with adapters?
Forum: Illumina/Solexa 02-12-2018, 03:47 PM
Replies: 5
Views: 1,168
Posted By hoytpr
You're right, that does resemble over...

You're right, that does resemble over amplification. I was going to run the samples on a denaturing gel but that protocol says not to. Can it be fixed? I think the protocol was mostly custom. I'm not...
Forum: Illumina/Solexa 02-12-2018, 02:18 PM
Replies: 5
Views: 1,168
Posted By hoytpr
Small RNA library with 200bp extra peak

I just looked at our new small RNA library submitted by a respected lab on the Bioanalyzer. The small RNA library should be about 178, and it's there, but most of the samples have a broader but...
Forum: Illumina/Solexa 02-09-2018, 02:40 PM
Replies: 6
Views: 1,040
Posted By hoytpr
Just to put a tack in this thread. ...

Just to put a tack in this thread.
Communication in cores is always a key to success. The client created a perfectly compatible small RNA Illumina library. They just didn't know it.

Edit: They...
Forum: Illumina/Solexa 02-08-2018, 12:14 PM
Replies: 6
Views: 1,040
Posted By hoytpr
Hadn't thought of custom primer spike-ins. What a...

Hadn't thought of custom primer spike-ins. What a cool idea!

Just add in the same number of pMoles... as long as one end of the library attaches to the flowcell, and the custom primers make it to...
Forum: Illumina/Solexa 02-08-2018, 07:45 AM
Replies: 6
Views: 1,040
Posted By hoytpr
Hi GW, Yes I mean cBot (:o). But after digging...

Hi GW,
Yes I mean cBot (:o). But after digging deep in the protocol I saw this:

# During cluster generation on the cBot, it is necessary to select
the ‘‘SR_TubeStripHyb’’ recipe because the...
Forum: Illumina/Solexa 02-06-2018, 02:05 PM
Replies: 6
Views: 1,040
Posted By hoytpr
PARE sequencing; single-read cluster generation

We got a request to do PARE sequencing (J. Zhai et al. / Methods 67 (2014) 84–90, http://dx.doi.org/10.1016/j.ymeth.2013.06.025) but someone correct me if I'm wrong.

The "single-read cluster...
Forum: Illumina/Solexa 01-10-2018, 11:20 AM
Replies: 13
Views: 2,929
Posted By hoytpr
Turnaround is really important. The market for...

Turnaround is really important. The market for these machines is the ~$100 per bacterial genome smaller groups. 1.2 Gbp is about 8-10 bacteria at 30X coverage. If the libraries are cheap enough...
Forum: Illumina/Solexa 01-10-2018, 05:10 AM
Replies: 2
Views: 742
Posted By hoytpr
Yes, and it was confirmed yesterday to be a bug...

Yes, and it was confirmed yesterday to be a bug in IEM 1.14 and 1.15 (just released), but not in 1.11. I have not tested 1.12, or 1.13.
Showing results 1 to 25 of 61

 


All times are GMT -8. The time now is 03:42 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2019, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO