SEQanswers

Go Back   SEQanswers > Search Forums


Showing results 1 to 25 of 56
Search took 0.00 seconds.
Search: Posts Made By: DonDolowy
Forum: Bioinformatics 11-04-2014, 05:34 AM
Replies: 46
Views: 17,269
Posted By DonDolowy
Thanks a lot for the help, guys! Working here...

Thanks a lot for the help, guys! Working here again.
Forum: Bioinformatics 10-28-2014, 07:25 AM
Replies: 46
Views: 17,269
Posted By DonDolowy
I have the same problem and the path is correct. ...

I have the same problem and the path is correct.

/Update: Thought it was related to Yosemite, as apparently there are some issues with Rstudio and Yosemite getting the path right. Tried to boot up...
Forum: RNA Sequencing 04-25-2014, 11:32 AM
Replies: 1
Views: 1,174
Posted By DonDolowy
Taqman and RNA-seq do not correlate

Dear all,

some time ago we did Taqman for a couple of candidate genes on some human samples, that showed downregulation of these genes. We therefore decided to go for a low-depth RNA-seq on these...
Forum: Bioinformatics 04-13-2014, 01:47 AM
Replies: 3
Views: 1,981
Posted By DonDolowy
Thanks a lot, guys. Very helpful!

Thanks a lot, guys. Very helpful!
Forum: Bioinformatics 04-11-2014, 11:38 PM
Replies: 3
Views: 1,981
Posted By DonDolowy
Generation of 192 barcode sequences that are furthest apart

Dear all,

we are working on implementing MARS-seq in our lab and for this purpose we would need to generate unique barcodes (6 bases barcode). We would like to scramble the X's (see below) to...
Forum: Bioinformatics 04-03-2014, 01:25 AM
Replies: 4
Views: 2,052
Posted By DonDolowy
If I want to compare my RNA-seq with publicly...

If I want to compare my RNA-seq with publicly available microarray datasets, I tend to work with fold expression instead of actual expression values. That should make the direct comparison legit.
Forum: Bioinformatics 03-25-2014, 06:31 AM
Replies: 3
Views: 1,480
Posted By DonDolowy
Correlating/clustering RNA-seq with ChIP-seq data?

Hi everyone,

I have RNA-seq data from adipocytes, where a certain gene, that is involved in heterochromatin, is knocked out. I would like to correlate/cluster this dataset with another dataset,...
Forum: Bioinformatics 03-05-2014, 05:06 AM
Replies: 3
Views: 1,712
Posted By DonDolowy
Yep, check out http://www.broadinstitute.org/gsea/

Yep, check out http://www.broadinstitute.org/gsea/
Forum: Bioinformatics 03-04-2014, 10:05 AM
Replies: 3
Views: 1,712
Posted By DonDolowy
Tjenar tjenar, you could either run GSEA on...

Tjenar tjenar,

you could either run GSEA on the dataset to see what gene sets are enriched in your conditions or you could go for GO analysis on your differentially expressed genes.
Forum: General 03-02-2014, 11:51 PM
Replies: 7
Views: 3,497
Posted By DonDolowy
I would definitely sequence at lower depth and...

I would definitely sequence at lower depth and sequence all of them for the same amount of money as if you were to pool them and sequence deeper (taken that you are just interested in gene...
Forum: Sample Prep / Library Generation 02-26-2014, 11:29 PM
Replies: 2
Views: 3,234
Posted By DonDolowy
MARS-seq (Parallel Single-Cell RNA-seq)?

Hi guys,

recently, a very interesting paper got published in Science how to do single-cell RNA-seq using a three-level barcoding system.

http://www.sciencemag.org/content/343/6172/776.abstract...
Forum: General 02-12-2014, 10:00 PM
Replies: 7
Views: 1,938
Posted By DonDolowy
In our case it does include library prep. We just...

In our case it does include library prep. We just hand over the RNA.
Forum: General 02-12-2014, 12:28 PM
Replies: 7
Views: 1,938
Posted By DonDolowy
We pay about 125€ per sample 2x50bp PE @ 15 mio...

We pay about 125€ per sample 2x50bp PE @ 15 mio reads. Does not get much cheaper than that!
Forum: Epigenetics 10-30-2013, 08:40 AM
Replies: 17
Views: 11,779
Posted By DonDolowy
Finally, it seems like it is working now! ...

Finally, it seems like it is working now!

Using a heat block or water bath instead of thermo cycler seems to do the trick. Incredible :)
No big changes between Fermentas and NEB's MspI enzyme.
Forum: Epigenetics 10-29-2013, 10:50 PM
Replies: 17
Views: 11,779
Posted By DonDolowy
That gel looks great. Far from what I am getting!...

That gel looks great. Far from what I am getting!
How do you extract your DNA? And are you also using MspI from NEB?
Forum: Epigenetics 10-29-2013, 07:38 AM
Replies: 17
Views: 11,779
Posted By DonDolowy
I now performed another MspI digestion on...

I now performed another MspI digestion on commercial available mouse gDNA.

2ug DNA
5uL Cutsmart buffer
1.5uL MspI enzyme
------
Total volume 50uL. 37C, 2hours.

Followed by cleanup using...
Forum: Epigenetics 10-28-2013, 09:19 AM
Replies: 17
Views: 11,779
Posted By DonDolowy
Hi C.R, thanks for the quick reply. The DNA I am...

Hi C.R, thanks for the quick reply. The DNA I am working on now is completely pure, since it is commercial DNA bought only for optimization to exclude potential problems with impure DNA.
I have also...
Forum: Epigenetics 10-28-2013, 09:03 AM
Replies: 17
Views: 11,779
Posted By DonDolowy
Hey again, I have now tried several times to...

Hey again, I have now tried several times to digest gDNA with MspI but I never get fragments in the low range (>500bp).
I have tried MspI enzyme from both NEB and Fermentas, which I have tested on...
Forum: Epigenetics 09-12-2013, 04:05 AM
Replies: 17
Views: 11,779
Posted By DonDolowy
MspI digestion - how is it supposed to look?

Hi all,

I am trying to prepare RRBS-seq library, but have some questions regarding the MspI digestion. I find my results a bit odd, since not much is digested and I have not been able to find a...
Forum: Bioinformatics 08-27-2013, 11:38 PM
Replies: 8
Views: 7,052
Posted By DonDolowy
I just find it odd that if you download a certain...

I just find it odd that if you download a certain iGenome "package" (e.g. UCSC mm9) that then the genome.fa and genes.gtf do not correspond and you get this error.

Personally, I have just removed...
Forum: Bioinformatics 08-12-2013, 09:43 AM
Replies: 10
Views: 4,841
Posted By DonDolowy
Thanks for all your input so far. Anders,...

Thanks for all your input so far.

Anders, you are absolutely right. I forgot to mention that the values are first normalized to TBP and then normalized to Ctrl group, which is the first black...
Forum: Bioinformatics 08-12-2013, 08:00 AM
Replies: 10
Views: 4,841
Posted By DonDolowy
Samples were 8-plexed giving us 25-30 million...

Samples were 8-plexed giving us 25-30 million mapped reads. This should be more than sufficient for gene expression analysis. We are not interested in splicing events etc, but use RNAseq as an...
Forum: Bioinformatics 08-12-2013, 07:14 AM
Replies: 10
Views: 4,841
Posted By DonDolowy
RNA-seq analysis is not consistent with qPCR results

Dear all,

I used to be of the opinion that RNAseq results do not need to be verified by qPCR, but to satisfy future reviewers I sat out to verify the genes of interest by qPCR. To my big surprise,...
Forum: Bioinformatics 08-07-2013, 09:06 AM
Replies: 3
Views: 2,411
Posted By DonDolowy
You can run GSEA to identify enriched gene sets...

You can run GSEA to identify enriched gene sets in (here you submit a matrix containing expression values for all genes in between your two conditions).
If you want to study only the DEGs, then you...
Forum: Bioinformatics 08-07-2013, 12:15 AM
Replies: 4
Views: 1,467
Posted By DonDolowy
I am using UCSC mm9 genes.gtf from iGenome. ...

I am using UCSC mm9 genes.gtf from iGenome.

I imagine I did something like:
Showing results 1 to 25 of 56

 


All times are GMT -8. The time now is 06:14 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2020, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO