FPKM = Fragments Per Kilobase Million FPKM is very similar to RPKM. RPKM was made for single-end RNASEQ, where every read corresponded to a single fragment that was sequenced. FPKM was made for paired-end RNA-seq. TPM = Transcripts Per Million (Sum of all TPM in samples is the same) TPM is very similar to RPKM and FPKM.

• TPM — This is salmon’s estimate of the relative abundance of this transcript in units of Transcripts Per Million (TPM). TPM is the recommended relative abundance measure to use for downstream analysis. • NumReads — This is salmon’s estimate of the number of reads mapping to each transcript that was quantified.

Practical Guide to Interpreting RNA-seq Data Skyler Kuhn1,2 Mayank Tandon1,2 1. CCR Collaborative Bioinformatics Resource (CCBR), Center for Cancer Research, NCI

• Use Salmon to “align” to Gencode transcriptome and generate both quant files (Read Count and TPM per transcript) for both transcripts and genes. Additionally, gather data from optional bootstrap for subsequent use in Sleuth DEG program. This stage designed to run with separate node for each sample.