SEQanswers

Go Back   SEQanswers > Search Forums


Showing results 1 to 17 of 17
Search took 0.00 seconds.
Search: Posts Made By: sbarberan
Forum: Sample Prep / Library Generation 06-08-2018, 02:54 PM
Replies: 270
Views: 107,692
Posted By sbarberan
Talking about which cells are used, is there...

Talking about which cells are used, is there anyone here with experience with single-cell experiments with MCF7?

Also, Simeone78, when you say play around with the lysis buffer, what exactly do...
Forum: Bioinformatics 03-26-2018, 01:39 PM
Replies: 1
Views: 422
Posted By sbarberan
Hi Ina, I use samtools and picard-tools to...

Hi Ina,

I use samtools and picard-tools to get the counts for miRNAs.

Here is what I would do for bowtie2 alignment to miRBase
bowtie2 -L8 --local -x miRBase-mature-hsa-index -U FILE.fastq -S...
Forum: Core Facilities 03-14-2018, 09:49 PM
Replies: 7
Views: 2,401
Posted By sbarberan
Hi Phillip, Really interesting points about...

Hi Phillip,

Really interesting points about purification.

You mention that using the RNA chip allows you to see primers/dimers even if they are annealed to full length library molecules; and...
Forum: Illumina/Solexa 02-13-2018, 11:22 AM
Replies: 5
Views: 944
Posted By sbarberan
You are probably right that qPCR will be the only...

You are probably right that qPCR will be the only way to quantify with 100% accuracy, but generally we do not use qPCR and even with libraries with bulged products we manage to quantify only with...
Forum: Illumina/Solexa 02-12-2018, 05:54 PM
Replies: 5
Views: 944
Posted By sbarberan
Bulged products are not a problem for sequencing,...

Bulged products are not a problem for sequencing, you only see them with gels or bioanalyzer. For sequencing you denature the libraries anyways. So there is nothing to fix, just that they probably...
Forum: Illumina/Solexa 02-12-2018, 04:05 PM
Replies: 5
Views: 944
Posted By sbarberan
That to me looks like over-amplification, we have...

That to me looks like over-amplification, we have seen 'bulged' products that run at 200-250bp. See page 9 of this protocol from the Mello lab (http://www.lsi.umich.edu/files/SmallRNACloning.pdf)
...
Forum: Sample Prep / Library Generation 02-08-2018, 02:46 PM
Replies: 3
Views: 1,165
Posted By sbarberan
I also was wondering if there are any users with...

I also was wondering if there are any users with experience with single-cell printing (http://www.cytena.com/SCP.html) and how this could compare to FACS sorting. Looks to me that cells undergo less...
Forum: Sample Prep / Library Generation 02-08-2018, 02:10 PM
Replies: 3
Views: 1,165
Posted By sbarberan
Thanks a lot Simone! We want to use FACS...

Thanks a lot Simone!

We want to use FACS sorting, and we are only looking for an instrument to reliably prepare 1/2 or 1/4 size libraries to reduce cost.

Cheers,
sergio
Forum: Sample Prep / Library Generation 01-26-2018, 11:16 AM
Replies: 3
Views: 1,165
Posted By sbarberan
Single-cell libraries miniaturization

Any advice from single-cell experts about the best liquid handler for miniaturization of libraries?

We are considering:
Mantis (Formulatrix)
Mosquito (ttplabtech)

Any comments about either...
Forum: Service Providers 01-16-2018, 12:07 PM
Replies: 2
Views: 2,348
Posted By sbarberan
I do not have experience with QIAseq targeted DNA...

I do not have experience with QIAseq targeted DNA panel. But can I ask what struggles do you have with cfDNA libraries?

How big is the panel that you are trying to profile?

Thanks!
Cheers,...
Forum: Bioinformatics 01-05-2018, 04:43 PM
Replies: 2
Views: 938
Posted By sbarberan
Hi Caroline, What I do is download the...

Hi Caroline,

What I do is download the sequences from miRBase and then change all 'U' to 'T' and then create the index with bowtie-build and then align.

Could it be that because you are not...
Forum: Sample Prep / Library Generation 12-14-2017, 09:52 PM
Replies: 5
Views: 1,123
Posted By sbarberan
I'm sure if you PAGE size-select the libraries...

I'm sure if you PAGE size-select the libraries will be fine to sequence.

For next time I highly recommend that you use a gel-free library preparation kit. I work for Somagenics, and we just...
Forum: Sample Prep / Library Generation 12-13-2017, 10:26 PM
Replies: 2
Views: 1,349
Posted By sbarberan
Did you find more information about this liquid...

Did you find more information about this liquid handler? We are also interested in this system, it does sounds like a perfect low-budget solution.
Forum: Sample Prep / Library Generation 12-13-2017, 10:21 PM
Replies: 5
Views: 1,123
Posted By sbarberan
How many cycles of PCR did you perform? We have...

How many cycles of PCR did you perform? We have seen a big peak ~280 when you over amplify libraries.

I think that these products at 280 could be 'bulge' products generated when you deplete...
Forum: Bioinformatics 12-13-2017, 10:14 PM
Replies: 1
Views: 1,067
Posted By sbarberan
To reduce problems with multiple mappings for...

To reduce problems with multiple mappings for miRNAs my approach is always align against miRBase first, and then the unaligned reads against the genome.

Why do you use bowtie and not bowtie2?
...
Forum: Bioinformatics 03-08-2017, 11:22 PM
Replies: 1
Views: 1,509
Posted By sbarberan
I use CAP-miRSeq that includes differential...

I use CAP-miRSeq that includes differential expression analysis with edgeR

http://bioinformaticstools.mayo.edu/research/cap-mirseq/
Forum: Bioinformatics 03-08-2017, 11:16 PM
Replies: 1
Views: 656
Posted By sbarberan
I found that when exploring new data CAP-miRSeq...

I found that when exploring new data CAP-miRSeq is really helpful:

http://bioinformaticstools.mayo.edu/research/cap-mirseq/
Showing results 1 to 17 of 17

 


All times are GMT -8. The time now is 11:10 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2018, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO