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Hi, If you have no replicate for your samples, then you can also use NOISeq package. The Pairwise Differential Expression Analysis (Without Replicates) functionality offers a strategy for analyzing RNA-seq datasets that do not have replicates. It is based on the software package NOISeq, which belongs to the Bioconductor project. NOISeq is a novel nonparametric approach for the identification for differentially expressed genes from RNA-Seq count data.