Moderator: swbarnes2

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Posts by swbarnes2

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Answer: A: Cluster sub-set analysis using Seurat
... Subsetting and rerunning the umap coords and clustering will separate out different things better. Personally, I cheat, and use Loupe. It's easy to use the mouse to draw categories, then export your categories into the metadata of your seurat object. But you have to have your data go through the ...
written 10 hours ago by swbarnes27.9k
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Answer: A: DESeq2: Getting results from multiple comparisons
... I hope you have a few replicates for each genotype and day combination. If you really only have 9 samples you can't do those comparisons. But if have replicates, make a new column that has genotype and days stuck together. Make that column name your design. You can specify whatever contrast you ...
written 11 hours ago by swbarnes27.9k
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Answer: A: fastqc result: whether trim off low quality reads or not
... It looks like bowtie2 will soft-clip to make an alignment happen. A phred score of 10 still means that the base is likely to be correct. The odds of wrong bases causing a read to align to the wrong place are pretty low. And if a read aligns nowhere, even after soft-clipping by an aligner, then ...
written 16 hours ago by swbarnes27.9k
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Comment: C: Why Salmon produces very different quantification results compared with featureC
... This is almost certainly it; salmon was intended to be smart about handling reads which have an ambiguous feature assignment, FeatureCounts was not. And uniquely mapping to a genome is not the same as uniquely mapping to one and only one gene feature. So including genes that multimap might not mak ...
written 19 hours ago by swbarnes27.9k
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Answer: A: Extracting Genes that contribute to 1st PC
... Now that you have code pZ <- prcomp(X_black, rank. = 3) #to get 1st 3 components So you are just using prcomp, which is part of base R. `pZ$rotation` will get you the loadings for the genes for each PC https://www.rdocumentation.org/packages/stats/versions/3.6.2/topics/prcomp ...
written 2 days ago by swbarnes27.9k
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Answer: A: DESeq2 Not Recognizing that my data set has been transformed when inputting. Hel
... I don't think you can import a vst transformed dataset like that. For one thing, when you run vst, you make a whole new DESeq Transform object out of it. It's not a slot that is added to the original dds object. I don't think that you can untransfrom vst counts, which is what you would need to fi ...
written 3 days ago by swbarnes27.9k
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Comment: C: Batch correction for one condition in DESeq2
... That command line doesn't normalize anything. Normalizing doesn't take your design into account at all. But ~ Genotype is the only design you should be using with that colData. ...
written 9 days ago by swbarnes27.9k
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Comment: C: Batch correction for one condition in DESeq2
... You cannot make use of a column where every single sample has the same value. There is no point in it being there. **You cannot get rid of or account for batch effect in the dataset you posted, because it is deeply confounded with genotype.** You can't make use of it, except as a guide to which g ...
written 9 days ago by swbarnes27.9k
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Answer: A: Batch correction for one condition in DESeq2
... If all your samples are primary, that doesn't belong in the ColData. Just drop it. The dates you have given are totally deeply confounded with your genotype. So you **have** to drop them too. If they really represent sequencing dates, then they aren't adding any technical artifacts. If they rep ...
written 9 days ago by swbarnes27.9k
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Comment: C: 10x Cell Ranger 'count' function error:
... It's looking for files with "R1" in the name, and not finding those. You can't proceed without them. ...
written 10 days ago by swbarnes27.9k

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