# Correct way to assemble reference transcriptome - what's --samples_file option in Trinity?

I have 101 samples from 9 tissues of a nonmodel species, coming either from SRA or sequenced by my lab. I want to generate a reference transcriptome with Trinity for further analysis. In order to do this, I downsampled the reads in order to have 20 to 30M reads for each tissue, for a total of 270M PE reads, which I combined in two files, one for left reads and one for right ones.

Now, I came across the --samples_file flag in Trinity and I'm not sure if I should use it instead of concatenating the transcriptomes. Any help on good practice on this?

--samples_file flag is for DE analysis, you can put info about biological replicates in that file. You say that you want to make a ref transcriptome, so you don't need a samples file for DE analysis.