User: wunderl

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wunderl20
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Posts by wunderl

<prev • 10 results • page 1 of 1 • next >
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Comment: C: DESeq2: Sample outlier patterns 'switching' after normalization
... Thanks, that makes sense. I can confirm from my data that the 'outlier' sample from the raw count plot had both the largest library size and highest variance. What fully convinced me was doing a simple normalization by library size. This ended up producing the same pattern of sample distances tha ...
written 3 months ago by wunderl20
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Answer: A: TMM Normalization: DESeqDataSet: some values in assay are negative
... You are likely getting this error because of `log=TRUE` in your call to `cpm()`. The `log` of anything less than 1 will be negative, so if after normalization you end up with any fractional counts they will produce a negative number when you take the `log`. This issue also occurs with DESeq2's `rl ...
written 4 months ago by wunderl20
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Comment: C: TMM Normalization: DESeqDataSet: some values in assay are negative
... If you want to truly compare pipeline performance, you should let each method do the normalization it's own way. Each method has been designed with its own philosophy and underlying assumptions, so mixing parts of one method with another is likely going to give you sub-optimal performance. (or, if ...
written 4 months ago by wunderl20
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DESeq2: Sample outlier patterns 'switching' after normalization
... ::Edit:: The embedded images are not formatting well, so I have changed them to external links. I have been following the [DESeq2 vignette][1] for bulk RNA-Seq data and got the to section where it is recommended you make a [heatmap of the euclidian distances between samples][2]. In addition to the ...
normalization deseq2 written 4 months ago by wunderl20 • updated 4 months ago by Michael Love26k
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Comment: C: Batch Correction: DESeq2 Linear Model vs Batch-Correction-Specific Algorithms (R
... Thank you for the elaboration. I didn't realize `removeBatchEffect` operated at the log-transformed level, I can see now why it is distinct from the suite of tools `DESeq2` provides. The point about GC bias was very interesting as well. I am very new to this sort of space so I really appreciate th ...
written 5 months ago by wunderl20
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Answer: A: DESeq2 GLM modelling and Wald's test
... I am new to `DESeq2` myself so take this answer with a grain of salt. I was initially confused by the difference between a `Wald test` and a `t-test` as well. To explain why a Wald test is used we first need to explain why we are fitting a generalized linear model (`GLM`). A **lot** is going on ...
written 5 months ago by wunderl20
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Comment: C: Batch Correction: DESeq2 Linear Model vs Batch-Correction-Specific Algorithms (R
... Thanks Michael, I really appreciate the help!! I will look into Salmon. So far my pipeline has been `STAR` --> `featureCounts` --> `DESeq2`. Would you say that not correcting for GC bias is a significant oversight? Also, is there a reason `DESeq2` does not have anything analogous to `limma ...
written 5 months ago by wunderl20
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All Pairwise Comparisons with DESeq2 While Controlling for Batch Effects: Does Reference Level Matter?
... I have the following design matrix: ``` batch treatment A 1 A 2 A 3 B 1 B 2 B 3 C 1 C 2 C 3 > batch [1] A A A B B B C C C Levels: A B C > treatment [1] 1 2 3 1 2 3 1 2 3 Levels: 1 2 3 ``` I wo ...
deseq2 differential expression design batch correction written 6 months ago by wunderl20 • updated 5 months ago by Michael Love26k
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Batch Correction: DESeq2 Linear Model vs Batch-Correction-Specific Algorithms (RUV, ComBat, ect.)
... I am analyzing bulk RNA data from neurons derived from iPSC cells. The experimental design matrix is as follows: | Lineage |     | Method ------------- | ------------ A | |1 A | | 2 A | | 3 B | | 1 B | | 2 B | | 3 C | | 1 C | | 2 C | | 3 Here `lineage` indicates what iPSC line the neuronal ...
deseq2 differential expression batch correction written 6 months ago by wunderl20 • updated 5 months ago by Michael Love26k
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featureCounts Does Not Correctly Parse File Names
... When using Rsubread v 1.28.1 it sucessfully parsed my input file names by removing the file path and just keeping the names. As of the current version, however, the names contain the entire file path, this makes for very long and ugly column names. Example ---- Setup --- files: /path/to/file/here/ ...
rsubread featurecounts written 6 months ago by wunderl20 • updated 6 months ago by Gordon Smyth39k

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