SEQanswers

Go Back   SEQanswers > Search Forums


Showing results 1 to 18 of 18
Search took 0.00 seconds.
Search: Posts Made By: Rosmano
Forum: Sample Prep / Library Generation 08-20-2019, 01:47 AM
Replies: 15
Views: 2,637
Posted By Rosmano
Yup, I got everything working by using solely the...

Yup, I got everything working by using solely the NEBNext master mix. :D

Now, I'm just wondering about performing bead clean-up... I've decided to do a Fragmet analyzer before doing size...
Forum: Sample Prep / Library Generation 08-14-2019, 01:48 AM
Replies: 9
Views: 5,641
Posted By Rosmano
I am doing the qPCR quantification with Power...

I am doing the qPCR quantification with Power SYBR Green PCR Master Mix from Thermo Fisher instead of the NEB Next HF Master Mix.

Does anyone know if this might impact the qPCR quantification?
Forum: Sample Prep / Library Generation 08-14-2019, 01:10 AM
Replies: 15
Views: 2,637
Posted By Rosmano
By the way, I am doing the qPCR quantification...

By the way, I am doing the qPCR quantification with Power SYBR Green PCR Master Mix from Thermo Fisher instead of the NEB Next HF Master Mix.

Does anyone know if this might impact the qPCR...
Forum: Sample Prep / Library Generation 08-12-2019, 08:53 AM
Replies: 15
Views: 2,637
Posted By Rosmano
Well, after analyzing my qPCR results again, what...

Well, after analyzing my qPCR results again, what I have to say is that the PCR looks awful but there's something there. Unfortunately, by the end of the 20 cycles I didn't hit a plateau stage and...
Forum: Sample Prep / Library Generation 08-10-2019, 08:15 AM
Replies: 15
Views: 2,637
Posted By Rosmano
What protease inhibitor did you use in your...

What protease inhibitor did you use in your protocol? I adopted the Omni-ATAC protocol but there's no qPCR amplification, even though I have DNA (7.6ng/uL obtained from 50,000 thymocytes).

I don't...
Forum: Sample Prep / Library Generation 08-10-2019, 08:07 AM
Replies: 9
Views: 5,641
Posted By Rosmano
Those two bands with less than 100bps are adapter...

Those two bands with less than 100bps are adapter dimers.

I am having problems in the step of pre-amplification.
After pre-amplification I get a DNA quantity of 7.6ng/uL from 50,000 total...
Forum: Sample Prep / Library Generation 04-03-2019, 02:04 AM
Replies: 1
Views: 806
Posted By Rosmano
As an update, I contacted Illumina and they said...

As an update, I contacted Illumina and they said they haven't text the Nextera DNA flex kit for ATAC-seq.

Or if anyone has any suggestion of an alternative kit that I can use I would welcome such...
Forum: Sample Prep / Library Generation 04-03-2019, 01:30 AM
Replies: 1
Views: 806
Posted By Rosmano
Unable to do library amplification using the new Nextera DNA Flex Library Prep kit

Hello everyone,

I am trying to do a library of FACS sorted T-cells using the new Nextera DNA Flex Library Prep kit to perform ATAC-seq.
We are using adapters ordered from Sigma that have the same...
Forum: Sample Prep / Library Generation 04-03-2019, 01:16 AM
Replies: 15
Views: 2,637
Posted By Rosmano
Hey, did you use the new Nextera DNA flex Library...

Hey, did you use the new Nextera DNA flex Library Prep kit?
I've been having similar problems to you but nothing I do seems to work.
Forum: Sample Prep / Library Generation 02-05-2013, 08:12 AM
Replies: 10
Views: 3,738
Posted By Rosmano
Hello! Thanks everyone to the replies....

Hello!

Thanks everyone to the replies. Well, we continued to troubleshoot the protocol and managed to get rid of that shoulder by doing two things:

1 - Do an Ampure XP beads purification with...
Forum: Illumina/Solexa 01-14-2013, 05:40 AM
Replies: 0
Views: 789
Posted By Rosmano
What is this shoulder in the BA profile?

Hey,

I am a newbie who has been trying to build ChIP libraries using the TruSeq ChIP Sample preparation kit.
After doing protocol optimization we decided to use MinElute columns for pre-PCR...
Forum: Sample Prep / Library Generation 01-14-2013, 02:00 AM
Replies: 10
Views: 3,738
Posted By Rosmano
So, I suppose that these Bioanalyzer curves don't...

So, I suppose that these Bioanalyzer curves don't have any issue?
Could this shoulder be derived from Y-shaped adapters?

I would like ot know if that shoulder may interfere with sequencing, since...
Forum: Sample Prep / Library Generation 01-11-2013, 03:04 PM
Replies: 10
Views: 3,738
Posted By Rosmano
I wouldn't mind to try that, but if possible in...

I wouldn't mind to try that, but if possible in the short term I would like to understand what issues may this shoulder on samples 2 and 4 cause.
Why it may be happening and ways to avoid while...
Forum: Sample Prep / Library Generation 01-11-2013, 10:44 AM
Replies: 10
Views: 3,738
Posted By Rosmano
What is this BA shoulder?

Hey,

I am a newbie who has been trying to build ChIP libraries using the TruSeq ChIP Sample preparation kit.
After doing protocol optimization we decided to use MinElute columns for pre-PCR...
Forum: Sample Prep / Library Generation 06-20-2012, 06:04 AM
Replies: 3
Views: 3,417
Posted By Rosmano
I shear my DNA by sonication. I repeated the...

I shear my DNA by sonication.
I repeated the PCR amplification (16 cycles this time) and the Ampure beads purification step with the other half of my samples. However, this time I changed the ratio...
Forum: Sample Prep / Library Generation 06-13-2012, 04:15 AM
Replies: 3
Views: 3,417
Posted By Rosmano
So what's the size of the adapter-dimer peaks...

So what's the size of the adapter-dimer peaks that make a sample bad for sequencing?
Is it when it is roughly the same size of the sample?
Forum: Sample Prep / Library Generation 06-08-2012, 08:08 AM
Replies: 3
Views: 3,417
Posted By Rosmano
Bioanalyzer results - adapter dimers in ChIP-seq library samples

Hello all,

I've recently found this forum and wanted to ask for feedback about the quality of the attached ChIP-seq libraries.

The relevant information is:
Used Bioo Scientific NextFlex...
Forum: Introductions 06-08-2012, 05:09 AM
Replies: 0
Views: 674
Posted By Rosmano
Greetings from Portugal

Hello everybody!

I am Rosmano, a PhD student in Portugal. My project is about the characterization of the transcriptional network of a certain transcription factor involved in neural stem cell...
Showing results 1 to 18 of 18

 


All times are GMT -8. The time now is 06:53 PM.


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