I am doing gene expression study for 2 different treatments in order to comparing them and i follow this methods:
RNA extraction,sequencing by illumina ,QC, trimmomated, assemblu uding de no vo , CEGMA, then i start Differential Expression Analysis Using a Trinity Assembly in command line following this steps:
Alignment-based abundance estimation methods
Build Transcript and Gene Expression Matrices
Counting Numbers of Expressed Transcripts or Genes(no biological replications)
Now i want run Extracting and clustering differential expressed transcripts but not sure should i do TMM normalisation before or no ?!! if yes How can i do TMM normalisation using trinity in commandline?!!