PCA plot (Suggestions Needed)
1
1
Entering edit mode
6 months ago
Aaliya ▴ 10

Hello, I have 8 samples (4 WT and 4 mutants). I have performed bulk RNA seq on these (Kallisto, and then DESeq2) I am attaching my PCA plot, I was expecting the wild type to cluster together and all the mutants to be together, but this was not the result. Does it indicate that my samples have a lot of variance?

Can anyone suggest to me any other method/plot that can represent my WT vs mutant samples?

I would really appreciate it.

enter image description here

DESeq2 PCAplot • 1.1k views
ADD COMMENT
0
Entering edit mode

How much variance were explained by PC1 and PC2? It doesn't look like that the largest source of variation came from the genotype difference.

ADD REPLY
0
Entering edit mode

Cross posted here (including the code used to make up the counts):

https://support.bioconductor.org/p/9157911/#9157959

ADD REPLY
2
Entering edit mode
6 months ago
Jeremy ▴ 930

Right now, the WT samples are clustering together, but you have one mutant sample off by itself, which could be an outlier. How did you scale the data? If you used Z-scores, I would suggest trying center log ratios.

Besides PCA, you could also try hierarchical clustering.

ADD COMMENT
2
Entering edit mode

Everything that Jeremy said, plus that there may be no data manipulation that will give you the clusters you expected. Sometimes there are outliers in the data, and that won't change no matter how much we torture the data.

ADD REPLY
1
Entering edit mode

You could also try kernel PCA, which is a non-linear form of PCA, although this will be more time-consuming.

ADD REPLY
1
Entering edit mode

Even if this was real data, the tiny numbers for the PC1 and PC2 indicate that the differences between the samples are all absolutely tiny.

ADD REPLY
0
Entering edit mode

I have actually used DESeq2 data.

ADD REPLY
1
Entering edit mode

The code you already posted in the bioconductor board had simulated data, and the PCA image is the same.

ADD REPLY

Login before adding your answer.

Traffic: 1574 users visited in the last hour
Help About
FAQ
Access RSS
API
Stats

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.

Powered by the version 2.3.6