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?


1 Answer 1


--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.

Why do you down sample? Are you not afraid you will not have enough reads to have a complete transcriptome now?

  • $\begingroup$ Thanks, it's what I was thinking. // I have 270 million reads, I think it's enough. Using all transcriptomes means having >750 million reads, I don't have the computational resources to assemble it. $\endgroup$ Jun 25, 2018 at 13:45
  • $\begingroup$ Okay, I understood 270 PE reads... $\endgroup$
    – benn
    Jun 25, 2018 at 13:53
  • $\begingroup$ Yes, I actually wrote 270 PE reads, my bad. $\endgroup$ Jun 25, 2018 at 13:54

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