I have some questions regarding RNA seq analysis if you can suggest anything it will help me a lot.
- I am currently normalizing RNA seq data for comparing genes expression within and between samples. Which normalization method would you recommend for this type of analysis? FPKM, TPM, TMM? Also I want to make a heatmap to see genes expressed in different conditions. Do you think transforming normalized data (like log2, z-score) is a good idea for this?
- Also I want to build a co expression network, I am just wondering if normalization like FPKM, TPM, TMM has any influence on building a coexpression network?
- Another thing I want to do is use Pearson correlation but I am confused if it will only find linear relationships among the normally distributed data. But normalization methods do not assume that counts to be normally distributed. So, Is it a bad idea to find pairwise coexpression of genes using pearson correlation? If so, which method would you recommend is reliable for building coexpression networks?
Please help me with this, I will be very grateful to you
Thanks in advance