Hello,
I am analyzing my small RNA seq data which contains 6 samples in total and includes two controls and 3 different treatments. In addition, one treatment has two different time points, so my controls also have two different time points. My coldata looks sth like this:
<caption>colData</caption>| condition | type | |
|---|---|---|
| sample 1 |
treatment 1 |
2 h |
|
sample 2 |
treatment 1 | 4 h |
| sample 3 |
untreated |
2 h |
| sample 4 |
untreated |
4 h |
| sample 5 |
treatment 2 |
2 h |
| sample 6 | treatment 3 |
2 h |
Now I am wondering how to analyze the data. Should I set a reference level for the first ctrl and run the full dataset and later repeat the analysis using the second ctrl as a reference level, or should I split the dataset to each individual one by one comparison, or should I omit setting a reference level and simply use the contrast argument in the results function? Or is this a case where the multi-factor design function should be applied?
Of course, I know that this kind of analysis is only for exploratory purposes, as I do not have biological replicates included.
In the end, I would like to come up with a heat map showing the treatment differences compared to their respective controls. As I do not want to show all genes, I will have to order in a way to show the same genes for each condition, but on the other hand, I want to show the relevant genes for each condition. Any suggestions how to order/ cluster?
One last question: On a windows machine I didn't have problems with plotting the resLFC in an MA plot using "plotMA", whereas my mac seemed to have problems. The Error message was: "Error from the generic function 'plotMA' defined in package 'BiocGenerics': no S4 method definition for argument 'resLFC' of class 'numeric' was found"
Thank you in advance!
