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Forum: RNA Sequencing 03-14-2019, 12:14 AM
Replies: 1
Views: 1,907
Posted By Noa
Optimal RNA prep from whole mouse tissue

I have an experiment planned in which RNA samples will be collected from total mouse tissue (eg whole kidney, whole brain, etc).
I plan to use Qiagen RNeasy kit for RNA prep and then do downstream...
Forum: RNA Sequencing 03-14-2019, 12:04 AM
Replies: 1
Views: 1,928
Posted By Noa
Optimal RNA preservation pre-sample prep over long time

I have an experiment planned in which RNA samples will be collected during a clinical study over the course of a year. I plan to use Qiagen RNeasy Micro kit for RNA prep

Which is the best option...
Forum: Bioinformatics 09-04-2014, 05:18 AM
Replies: 4
Views: 1,785
Posted By Noa
brilliant! thanks, saved me a ton of searching. ...

brilliant! thanks, saved me a ton of searching.
anyone know of other places to do this for non-cancer diseases??
Forum: Bioinformatics 09-04-2014, 05:06 AM
Replies: 4
Views: 1,785
Posted By Noa
Will try; thanks!

Will try; thanks!
Forum: Bioinformatics 09-04-2014, 04:23 AM
Replies: 4
Views: 1,785
Posted By Noa
How to search for a known mutation in WGS of disease?

I am working with researchers who have identified a gene mutation which significantly affects protein function.
They want to search all existing NGS (esp cancer) data to see whether this protein...
Forum: Bioinformatics 05-19-2014, 10:29 PM
Replies: 2
Views: 962
Posted By Noa
The genome is in scaffolds but reasonably...

The genome is in scaffolds but reasonably complete. I am not sure whether blast will be sensitive enough given the CG richness of the islands etc. Happy if anyone has any previous experience with...
Forum: Bioinformatics 05-19-2014, 04:44 AM
Replies: 2
Views: 962
Posted By Noa
comparing CpG islands between species

Hi all,
We are considering using chips developed for human CpG island detection for non model organism methylation analyses
Does anyone have useful insights into how best to compare CpG islands...
Forum: Sample Prep / Library Generation 03-31-2014, 01:33 AM
Replies: 0
Views: 13,259
Posted By Noa
Best way to make metagenomic library

We have several samples of DNA sampled from the ocean and want to make metagenomic libraries for a Illumina 150PE run. We are debating between doing overlapping reads or an insert size of 400-500bp....
Forum: Sample Prep / Library Generation 11-06-2013, 10:45 PM
Replies: 1
Views: 1,286
Posted By Noa
Werid RNA traces- bump above 28S - can I make libraries from this?

Hi all,
I have traces from a fragment analyzer (very similar to bioanalyzer; basically your capillary gel run) and am getting some weird traces. Some (well A2, A6 and A7 for example) have things...
Forum: Bioinformatics 10-24-2013, 12:39 AM
Replies: 2
Views: 1,364
Posted By Noa
Biostatistics help in planning experiment with downstream bioinfo in mind!

I have a complicated experimental system that is looking at changes in a region of the brain during learning. Past NGS has shown that, perhaps not surprisingly for brain tissue, there is a lot of...
Forum: Illumina/Solexa 10-22-2013, 10:07 PM
Replies: 2
Views: 1,868
Posted By Noa
Hi rebyl- I had done my trinity build on peaks...

Hi rebyl-
I had done my trinity build on peaks that were smears from 200-700 in the end. I had written the trinity rnaseq users group and brian haas answered me as copy-pasted below. The trinity...
Forum: Metagenomics 10-08-2013, 05:00 AM
Replies: 2
Views: 2,165
Posted By Noa
Sorry for the confusion! We have a WGS...

Sorry for the confusion!
We have a WGS metagenome; ignore the word amplicon here. I think they somehow tried to select for short fragments when the prepared the library but I will double check on...
Forum: Metagenomics 10-08-2013, 04:11 AM
Replies: 2
Views: 2,165
Posted By Noa
Best way to search for gene homologue in metagenomic dataset

Hi all,

This is my first foray into metagenomics so :

A collaborator has a metagenomic dataset (Illumina 100bpPE with ~200bp amplicons) and wants to search for her favorite gene in the dataset....
Forum: Bioinformatics 08-18-2013, 10:42 AM
Replies: 0
Views: 1,494
Posted By Noa
Possible bias from making library from edge of Covaris-sheared DNA?

Hi
We used a Covaris machine to shear BAC DNA that is supposed to go on a 2x250bp MiSeq run. The specifications used were as recommended by Covaris website:

Target BP (peak) 500:
Peak incident...
Forum: Illumina/Solexa 08-18-2013, 10:41 AM
Replies: 1
Views: 2,482
Posted By Noa
Making library from edge of sheared DNA smear - possible biases?

Hi
We used a Covaris machine to shear BAC DNA that is supposed to go on a 2x250bp MiSeq run. The specifications used were as recommended by Covaris website:

Target BP (peak) 500:
Peak incident...
Forum: Bioinformatics 08-05-2013, 11:57 PM
Replies: 0
Views: 1,646
Posted By Noa
Species comparison of number of hits by dc-megablast

Hi all,
I have a new transcriptome and am interested in comparing how many hits I get if I blast it to various similar sequences. I ran a dc-megablast against a few species of interest. Any...
Forum: Bioinformatics 07-04-2013, 03:45 AM
Replies: 2
Views: 1,748
Posted By Noa
Thanks for the clarification. Any idea what the...

Thanks for the clarification. Any idea what the differences are or how different the different annotations are? Is it worth running cuffdiff with the different annotations or would it all come out...
Forum: Bioinformatics 07-04-2013, 02:25 AM
Replies: 2
Views: 1,748
Posted By Noa
Compatability of UCSC NCBI and ENSEMBL genomes

I ran I ran Cuffdiff using an ENSEMBL genome (rat RGSC3.4) and want to look at my alignments using IGV
I tried to upload the genome and gtf file I downloaded from ENSEMBL to IGV but it is not...
Forum: Bioinformatics 07-02-2013, 10:50 PM
Replies: 0
Views: 1,088
Posted By Noa
Cuffdiff with biological replicates vs individual samples not adding up.

I am performing differential expression analysis of three different experimental conditions, each of which has 4 biological replicates, and was trying to do this using cuffdiff. We are really only...
Forum: Illumina/Solexa 05-22-2013, 12:16 PM
Replies: 14
Views: 8,245
Posted By Noa
So does anyone have a clue why you would use...

So does anyone have a clue why you would use different final NaOH concentrations to denature the same DNA? The libraries should be the same in the end regardless of whether they go on MiSeq or HiSeq,...
Forum: Illumina/Solexa 05-22-2013, 10:38 AM
Replies: 14
Views: 8,245
Posted By Noa
Has anyone ever used this protocol on a HiSeq...

Has anyone ever used this protocol on a HiSeq (and not a MiSeq)?
Thanks!
Forum: Illumina/Solexa 05-22-2013, 09:58 AM
Replies: 14
Views: 8,245
Posted By Noa
the protocol is from our genome center- Illumina...

the protocol is from our genome center- Illumina cBot user guide - (manual is Illumina part # 15006165 rev. K) from Oct 2012

(attached)

note it specifically says:

NOTE
If your application...
Forum: Illumina/Solexa 05-21-2013, 04:01 AM
Replies: 14
Views: 8,245
Posted By Noa
Are you also using MiSeq? Why is everyone using a...

Are you also using MiSeq? Why is everyone using a final of 0.1N NaOH? The protocol I have from Illumina uses a final of 0.05N (see below):

1 Combine the following volumes of template DNA and 0.1 N...
Forum: Illumina/Solexa 05-20-2013, 10:21 PM
Replies: 14
Views: 8,245
Posted By Noa
Thanks for taking the time to answer: Further...

Thanks for taking the time to answer: Further clarifications/questions:

(a) regarding the NaOH concentration- I am using the HiSeq 2500 - their protocol says to mix 10uL 0.1N NaOH with 10uL...
Forum: Illumina/Solexa 05-20-2013, 11:26 AM
Replies: 14
Views: 8,245
Posted By Noa
Preparation of low concentration libraries for hybridization

Hi,
I know this has been covered by many threads (I've read them all) but am looking for a few clarifications.

(1) I understand Illumina recommendations are such that you dont want to go past...
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