SEQanswers

Go Back   SEQanswers > Search Forums


Showing results 1 to 17 of 17
Search took 0.00 seconds.
Search: Posts Made By: FWOS
Forum: Sample Prep / Library Generation 07-18-2016, 11:41 AM
Replies: 1
Views: 1,199
Posted By FWOS
I've tried the kit with lower input from stranded...

I've tried the kit with lower input from stranded RNA library preps and it seems to be fine at low coverage (~40X). i'd probably try to stay above 150-200ng for the hybs if you're trying to cut down...
Forum: Illumina/Solexa 04-08-2015, 06:03 AM
Replies: 3
Views: 2,061
Posted By FWOS
I've seen the same thing. Some folks are...

I've seen the same thing. Some folks are doubting the adapters and adding double (5ul of 15uM index per reaction). From what i have tested it looks like ligation efficiency is much better with low...
Forum: Illumina/Solexa 01-13-2015, 09:57 AM
Replies: 53
Views: 87,241
Posted By FWOS
Pretty sure they will initially only support...

Pretty sure they will initially only support 2x125 in 3.5 days
Forum: General 12-09-2014, 09:39 AM
Replies: 16
Views: 4,166
Posted By FWOS
Hi Ganesh, I think it's as simple as this: ...

Hi Ganesh,

I think it's as simple as this:

RNA loves U

:eek:
Forum: Sample Prep / Library Generation 09-16-2014, 08:50 AM
Replies: 7
Views: 5,041
Posted By FWOS
We use the LE220 and can run 24 samples at a...

We use the LE220 and can run 24 samples at a time. The tubes are larger than regular shear tubes, so the adapter plate can only handle 24 per run. Not too bad to work with the tubes since they have...
Forum: Illumina/Solexa 09-16-2014, 08:43 AM
Replies: 4
Views: 3,398
Posted By FWOS
Thumbs up Ussually you'll want to HPLC purify for good...

Ussually you'll want to HPLC purify for good measure and I would ensure that your primers meet the following criteria -

Custom primers must:
Anneal to the P5 end of the library (refer to...
Forum: Sample Prep / Library Generation 09-09-2014, 07:43 AM
Replies: 7
Views: 5,041
Posted By FWOS
Hi, One of the reasons we moved forward with...

Hi,

One of the reasons we moved forward with Covaris was the higher yields for both DNA and RNA. (Compared to qiagen all prep FFPE; Kapa Express extract and 5 other kits we tested. The only...
Forum: Sample Prep / Library Generation 09-09-2014, 06:16 AM
Replies: 7
Views: 5,041
Posted By FWOS
I have been using the kit for about 6 months and...

I have been using the kit for about 6 months and it seems to work well. The protocol is fast and limits the amount of heat needed by only heating up for the ProtK treatment (compared to other preps...
Forum: Illumina/Solexa 05-17-2014, 09:15 AM
Replies: 8
Views: 7,471
Posted By FWOS
Hi, Just wanted to note that the Illumina...

Hi,

Just wanted to note that the Illumina nano and per free adapters are both Y adapters. The Y is created by improper base pairing at the 3' and 5' ends of the adapter, but that goes away once...
Forum: Bioinformatics 11-22-2013, 12:43 PM
Replies: 4
Views: 2,947
Posted By FWOS
All Illumina HiSeq reads start out lower. Mostly...

All Illumina HiSeq reads start out lower. Mostly this is improved after template generation (cycle 4). This is partially due to lower quality bases that result from the prep process, and it is also...
Forum: Sample Prep / Library Generation 11-18-2013, 11:56 AM
Replies: 2
Views: 3,198
Posted By FWOS
Thumbs up Hi Avo, I have been using the truseq...

Hi Avo,

I have been using the truseq PCR-free preps for a year now and have never run into issues with yield at the end. Assuming you are not losing a ton of material in the shear, the double...
Forum: Sample Prep / Library Generation 07-24-2013, 07:10 AM
Replies: 9
Views: 11,135
Posted By FWOS
Nextera Rapid fixed the issues I've had

The Nextera Rapid kit fixes issues with tagmentation efficiency using 50ng input gDNA. I have toyed with the number of PCR cycles required to yield sufficient .tagmented libraries for Exome prep...
Forum: Bioinformatics 04-09-2013, 05:53 AM
Replies: 25
Views: 6,383
Posted By FWOS
Library Type?

Hi Santos,

What kind of prep was done on these libraries? If the initial sequences are not diverse, you can see a wavy pattern in the first few bases. This happens with RNA seq libraries and can...
Forum: Bioinformatics 03-20-2013, 07:41 AM
Replies: 2
Views: 2,704
Posted By FWOS
Can be Normal

Hi Nico,

Your results seem normal to me. On average, One can expect to map 40 - 60% of the IP'd reads. I have seen less than 10% mapping back, which was actually due to salmon sperm blockers...
Forum: Illumina/Solexa 10-04-2012, 03:29 PM
Replies: 18
Views: 14,403
Posted By FWOS
Cbot denaturation

I am pretty sure that the cbot relies more on heat denaturation than on the NaOH base denaturation. So the base is just a failsafe. The first step in cbot cluster gen is an incubation around 98C. ...
Forum: Epigenetics 02-09-2012, 03:09 PM
Replies: 9
Views: 7,723
Posted By FWOS
Help with Quantification of ChIP Samples

Hi,
Thanks for the info and the protocol. I have some ChIP samples that I wanted to run through the Illumina sample prep protocol, but it seems like they can't be accurately quantified via the...
Forum: Bioinformatics 05-25-2011, 12:56 PM
Replies: 11
Views: 22,728
Posted By FWOS
Help Interpreting mRNA Seq Duplicate Sequence Plot FastQC

Hi All,

I recently noticed some strange trends in the duplicate sequence plots generated from a 2x50bp RNA sequencing experiment performed on an Illumina HiSeq. I understand that the libraries...
Showing results 1 to 17 of 17

 


All times are GMT -8. The time now is 02:25 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2021, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO