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Forum: Bioinformatics 12-08-2015, 07:33 PM
Replies: 24
Views: 10,883
Posted By N311V
Hi ewels, I've finally got time to run...

Hi ewels,

I've finally got time to run MultiQC but having trouble providing a list of directories to search. Do I need top put my fastqc and tophat results all in the same directory to produce a...
Forum: RNA Sequencing 11-24-2015, 02:42 PM
Replies: 5
Views: 2,277
Posted By N311V
The result seems reasonable. Is there any...

The result seems reasonable.

Is there any other reason you can think of that might cause systematic up regulation?

Were controls and treatments extracted at the same time? Sequenced at the same...
Forum: RNA Sequencing 11-18-2015, 11:05 AM
Replies: 5
Views: 2,277
Posted By N311V
I can't comment on your edgeR problem but maybe...

I can't comment on your edgeR problem but maybe try using limma and see if a different approach gives the same result.

Regarding 90% up-regulated, what are the differences between your cases and...
Forum: Sample Prep / Library Generation 11-15-2015, 04:17 PM
Replies: 7
Views: 2,036
Posted By N311V
I've got KAPA and NEBnext trail kits on the way,...

I've got KAPA and NEBnext trail kits on the way, yay! I'll post a comparison after I've used them.
Forum: Sample Prep / Library Generation 11-12-2015, 06:31 PM
Replies: 7
Views: 2,036
Posted By N311V
Awesome! Thanks nucacidhunter. It's a relief...

Awesome!

Thanks nucacidhunter. It's a relief not having to worry about buying the wrong kit.
Forum: Sample Prep / Library Generation 11-12-2015, 06:19 PM
Replies: 7
Views: 2,036
Posted By N311V
Hi necacidhunter, Thanks for the advice and...

Hi necacidhunter,

Thanks for the advice and yes we're considering in house prep as there are a lot to do.

Quantity of total RNA per sample won't be a problem and based on the current draft...
Forum: Bioinformatics 11-12-2015, 04:16 PM
Replies: 5
Views: 2,454
Posted By N311V
I've used bwa mem but not via that kit. Does...

I've used bwa mem but not via that kit.

Does running /r/bwa-kit/run-bwamem give you the available options?
Forum: Sample Prep / Library Generation 11-12-2015, 04:09 PM
Replies: 7
Views: 2,036
Posted By N311V
KAPA mRNA-seq kit, would you recommend it?

I'm considering use of KAPA Stranded mRNA-Seq Kits.

If anybody here is using it (or not but still has an opinion) I'd value your thoughts on the quality of library produced and it's ease of use.
...
Forum: Sample Prep / Library Generation 11-11-2015, 02:02 PM
Replies: 3
Views: 1,682
Posted By N311V
Hi James, Thanks for your advice. For...

Hi James,

Thanks for your advice.

For past experiments I've been involved in library prep was outsourced and that's the kit they used too. The results were also very good. However, cost wise I...
Forum: Bioinformatics 11-10-2015, 03:20 PM
Replies: 15
Views: 4,878
Posted By N311V
Hi Brian, Thanks for the reply and heads up...

Hi Brian,

Thanks for the reply and heads up regarding the HiSeq4000.

I have some V2 RNA-seq reads I'd like to run through CalcTrueQuality but they've been mapped with TopHat. Can I remap them...
Forum: Bioinformatics 11-09-2015, 02:29 PM
Replies: 15
Views: 4,878
Posted By N311V
Is this only a problem for the V1 kit?

Is this only a problem for the V1 kit?
Forum: Bioinformatics 11-09-2015, 12:08 PM
Replies: 24
Views: 10,883
Posted By N311V
That redefines what I mean by a lot also, I was...

That redefines what I mean by a lot also, I was talking about 142 samples! I'll hopefully get a chance to run it today.
Forum: Bioinformatics 11-08-2015, 06:43 PM
Replies: 1
Views: 1,042
Posted By N311V
Personally I think option a) is easier to...

Personally I think option a) is easier to interpret.

Given the genome as a background, there are 32 enriched GO terms in this set of differentially expressed genes.

If you've applied an...
Forum: Bioinformatics 11-08-2015, 06:14 PM
Replies: 24
Views: 10,883
Posted By N311V
Looks great! I have a heap of fastQC and Tophat...

Looks great! I have a heap of fastQC and Tophat files I can test it out on.
Forum: Sample Prep / Library Generation 11-08-2015, 04:58 PM
Replies: 3
Views: 1,682
Posted By N311V
Help choosing library prep for illumina

Hi All,

We have a large RNA-seq experiment planned (320 samples) for which I've been tasked with library prep. The amount and quality of RNA per sample should not be a problem as use of inbred...
Forum: Bioinformatics 10-28-2015, 03:43 PM
Replies: 7
Views: 4,233
Posted By N311V
It is also my understanding that mapping quality...

It is also my understanding that mapping quality in that case would be zero for both.

Is there an option to randomly keep one of the multiple mappings rather than discard all of them in bwa mem?
Forum: Bioinformatics 10-07-2015, 02:50 PM
Replies: 0
Views: 879
Posted By N311V
Multiplexing 60+ samples, will sampling/technical variance be too high?

Hi All,

The goal is differential expression analysis of a species of fly and I'd appreciate some opinions on multiplexing a large number of samples (60+) and sequencing several technical...
Forum: Bioinformatics 10-15-2014, 06:49 PM
Replies: 2
Views: 1,615
Posted By N311V
Thanks for the reply, it was very helpful. I...

Thanks for the reply, it was very helpful. I don't know how often genes will overlap on opposite strands for the species I'm working with but I'd be surprised if it doesn't happen a fair bit. So far...
Forum: Bioinformatics 10-15-2014, 05:13 PM
Replies: 2
Views: 1,615
Posted By N311V
mRNA sequencing - stranded versus not

Hi all,

We're in the early stages of planning an RNAseq experiment with two goals. One is eQTL and or differential expression analysis and the other is denovo assembly of the transcriptome....
Forum: Bioinformatics 07-31-2014, 03:58 PM
Replies: 8
Views: 2,308
Posted By N311V
With pretty much all of our HiSeq data we do see...

With pretty much all of our HiSeq data we do see a drop-off in quality towards the end of the second read but nothing that extreme. I tend to agree with previous comments that something has gone...
Forum: Bioinformatics 07-31-2014, 02:19 PM
Replies: 6
Views: 1,884
Posted By N311V
I did read somewhere on the GATK website that...

I did read somewhere on the GATK website that each sample needs a unique read group, sorry don't have a link right now. To keep track of lane perhaps you could use picard tools...
Forum: Bioinformatics 07-31-2014, 02:19 AM
Replies: 2
Views: 1,107
Posted By N311V
Was that ~11% of GEO projects from cell cultures...

Was that ~11% of GEO projects from cell cultures that had reads mapping to mycoplasma or ~11% of all GEO projects between 2012 and 2013 regardless of origin?
Forum: Bioinformatics 07-31-2014, 02:05 AM
Replies: 6
Views: 1,884
Posted By N311V
In regards to your first question, I do think...

In regards to your first question, I do think GATK UnifiedGenotyper would have treated each different read group as a different sample...
Forum: Bioinformatics 07-29-2014, 05:11 PM
Replies: 4
Views: 2,381
Posted By N311V
I've not worked with RNAseq data yet so I'm no...

I've not worked with RNAseq data yet so I'm no expert but my opinion from a statistical stand point is to agree with your reasoning.

I particularly dislike option 3 if you're interested in...
Forum: Bioinformatics 07-29-2014, 04:38 PM
Replies: 2
Views: 1,068
Posted By N311V
I'm not aware of a ready-made tool to do this job...

I'm not aware of a ready-made tool to do this job but it sounds like something you could do with Perl. I'm sure there are online tutorials you could use otherwise I found this book useful when first...
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