Forum: Sample Prep / Library Generation
05-17-2016, 09:49 AM
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Replies: 4
Views: 2,312
My main reason why not to use the spike-ins for...
My main reason why not to use the spike-ins for quantitative normalization is that I've never seen it demonstrated to work and that we are aware of biases that affect them - and these biases may vary...
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Forum: Bioinformatics
05-13-2016, 01:50 PM
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Replies: 4
Views: 2,421
I've spent a good deal of time working on the...
I've spent a good deal of time working on the ERCCs and have a few points to make in their defense;
Yes, polyA selection biases affect some ERCC controls. There is no reason to believe this is...
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Forum: Sample Prep / Library Generation
05-13-2016, 01:33 PM
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Replies: 4
Views: 2,312
A few points:
RUV really doesn't work well...
A few points:
RUV really doesn't work well with ERCCs; it doesn't account for the fact that the controls have built-in differences. If you do want to use RUV for this type of purpose, you'll need...
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Forum: Bioinformatics
06-29-2015, 01:30 PM
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Replies: 15
Views: 10,884
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Forum: RNA Sequencing
05-14-2015, 03:40 PM
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Replies: 1
Views: 1,255
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Forum: Bioinformatics
03-02-2015, 09:24 AM
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Replies: 15
Views: 11,995
In my opinion, ERCC spike-ins are uniquely...
In my opinion, ERCC spike-ins are uniquely situated to determine the mRNA:totalRNA ratio between samples, and are best used when you expect that the ratio is NOT equal. The Nature Biotech paper...
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Forum: RNA Sequencing
12-04-2014, 10:07 AM
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Replies: 3
Views: 2,425
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Forum: RNA Sequencing
12-02-2014, 12:01 PM
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Replies: 3
Views: 2,425
I have been working on a method to "unmix"...
I have been working on a method to "unmix" RNA-seq data from mixtures for awhile, and I think it may be of some help to your situation.
You would need to:
*Be able to collect a sample of your...
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Forum: Bioinformatics
11-24-2014, 05:24 PM
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Replies: 10
Views: 3,943
I did a quick test of this today and saw no...
I did a quick test of this today and saw no obvious difference (except for a uniformly higher FPKM in "standard") between standard cufflinks and using --no-effective-length, as regards the reported...
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Forum: Bioinformatics
11-18-2014, 11:07 AM
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Replies: 1
Views: 1,018
I have done some empirical mixture validation...
I have done some empirical mixture validation where I show that spike-ins do an excellent job of determining the different amounts of RNA per mixture component, assuming that you spike into a known...
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Forum: Bioinformatics
11-10-2014, 03:34 PM
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Replies: 3
Views: 1,943
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Forum: Illumina/Solexa
10-15-2014, 09:43 AM
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Replies: 1
Views: 1,099
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Forum: Illumina/Solexa
10-03-2014, 01:10 PM
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Replies: 2
Views: 1,379
State of the Art - RNAseq
This question really boils down to the following: If you were to do an RNA-seq experiment today (let's limit the sequencing to illumina) what protocols/kits would you use?
*For single-cell /...
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Forum: Sample Prep / Library Generation
07-16-2014, 10:28 AM
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Replies: 289
Views: 155,164
From the discussions i've had with C1 users,...
From the discussions i've had with C1 users, Spike-ins for single cell are unfortunately very cell-type specific - you want to maintain a 3-5% amount of external RNA to cellular RNA, but the amount...
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Forum: Bioinformatics
04-28-2014, 03:48 PM
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Replies: 4
Views: 2,488
Why are you using RPKM here?
I'm not too...
Why are you using RPKM here?
I'm not too familiar with your study, but I'd think that you want to be looking at windows of very specific width (around the TSSs, +/- N nt) and therefore will be...
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Forum: Bioinformatics
04-25-2014, 02:57 PM
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Replies: 1
Views: 1,024
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Forum: RNA Sequencing
04-21-2014, 11:22 AM
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Replies: 1
Views: 3,459
The practical difference between the mixes is...
The practical difference between the mixes is that most of the spike ins are at different concentrations between mixes.
You can use this difference to determine things - particularly detection...
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Forum: Bioinformatics
04-21-2014, 11:05 AM
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Replies: 2
Views: 1,796
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Forum: Bioinformatics
03-20-2014, 03:33 PM
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Replies: 15
Views: 6,921
Awhile ago I did an analysis that compared the...
Awhile ago I did an analysis that compared the list of genes called as "Differentially expressed" across a set of mixture samples by a number of different algorithms. These mixture samples had some...
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Forum: RNA Sequencing
01-21-2014, 10:40 AM
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Replies: 5
Views: 2,548
Illumina's human body map includes data for a...
Illumina's human body map includes data for a bunch of tissues (adipose adrenal brain breast colon heart kidney liver lung lymphnode ovary prostate muscle testes thyroid whiteblood) and has data out...
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Forum: Bioinformatics
01-07-2014, 10:16 AM
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Replies: 15
Views: 11,995
The DESeq size factors assume that most things...
The DESeq size factors assume that most things will be 1:1, so the 1:1 sub pool would be a good fit.
That said, it's data analysis so there's nothing stopping you from doing it both ways - I'd be...
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Forum: Bioinformatics
12-16-2013, 02:16 PM
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Replies: 2
Views: 2,981
Assa,
You should be concerned with the...
Assa,
You should be concerned with the effect of the differential spike-in between the treatment and control samples. This effect may be quite minor or more serious, depending on the percentage...
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Forum: Bioinformatics
10-22-2013, 09:35 AM
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Replies: 10
Views: 3,530
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Forum: General
09-19-2013, 12:33 PM
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Replies: 8
Views: 3,680
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Forum: RNA Sequencing
08-29-2013, 01:02 PM
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Replies: 11
Views: 13,198
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