Question: Unusual graph for Identifying HVGs from the normalized log-expression
0
gravatar for es.azadian
14 months ago by
es.azadian0
es.azadian0 wrote:

Hello to everyone,

I've followed the "A step-by-step workflow for low-level analysis of single-cell RNA-seq data with Bioconductor, V2" for my own data. The problem is that when I produce the graph for Variance of normalized log-expression values (using trendVar and decomposeVar functions), my plot (an ascending-linear shape in which all of dots are accumulated and correlated with the line of y=x, and it's not at all a bell-shaped graph) is basically different with the plot of workflow (Figure 10). Is it normal?!

I'm wondering if you could give me some hints.

 

 

workflows simplesinglecell • 269 views
ADD COMMENTlink modified 14 months ago by Aaron Lun25k • written 14 months ago by es.azadian0
Answer: Unusual graph for Identifying HVGs from the normalized log-expression
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gravatar for Aaron Lun
14 months ago by
Aaron Lun25k
Cambridge, United Kingdom
Aaron Lun25k wrote:

I'm not a telepath so these issues are very difficult to diagnose without code and plots (see the posting guide). Nonetheless, there are two obvious possibilities that come to mind.

  • You're computing the mean-variance trend on the raw counts instead of the log-transformed normalized counts. For Poisson-distributed counts, this should give a straight line.
  • You have run the code correctly but your counts are so low that you are effectively only visualizing the linear part of the plot. This would not be impossible for very low-coverage droplet-based data.

In any case, I would suggest following the updated instructions at http://bioconductor.org/packages/devel/workflows/html/simpleSingleCell.html rather than reading the article, as almost all of the code in the latter is out of date.

ADD COMMENTlink modified 14 months ago • written 14 months ago by Aaron Lun25k
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