Transcriptome analysis in D. discoideium (AX2) in different developmental stages

The goal of RNA-seq is to identify the differential expressed genes in the wild-type D.discoideum at different developmental stages (vegetative, streaming, mound and fruiting bodyies). Two biological replicates were assigned for each group and in total 8 groups were prepared for RNA-seq libraries with 500 ng mRNA as starting materials using NEXTflex Illumina qRNA-Seq Library Prep Kit (Bioo Scientific). We mapped about 20 - 30 million reads per sample to D. discoideum with bowtie2 workflow. The analysis pipeline started with pseudo-alignment of the reads using kallisto, followed by building an index and quantification process, which generated h5 files containing the raw binary files for preliminary differential analysis in sleuth. Differential analysis were conducted in EdgeR. The significant genes were deducted by computing NB exact genewise tests with corrected p < 0.05. Our results showed stage-specific gene expression profiles in different developmental stages. Overall design: Messenger RNA from vegetative, streaming, mound and fruiting body stages were extacted and cDNA libraries were generated for deep sequencing, in duplicate, using Nextseq500