Next Generation Sequencing for Transcriptomes Analysis for LAG-1 time course by auxin treatment.

Purpose: Ultilize Next Generation Sequencing to identify genes that are activated or repressed by LAG-1 (following auxin treatment to specifically deplete LAG-1 in C. elegans germline). The list of genes generated from RNA-seq identify primary and secondary targets of LAG-1 in C. elegans germline stem cells. Methods: RNA isolated from hand dissected C. elegans germline tumor (one day old adult animals) were used for RNA-seq library. Results: two genes (lst-1 and sygl-1) were identified as primary target for LAG-1 (2 hours auxin treatment); five additional genes are identified as secondary targets of LAG-1 based on their RNA accumulation depend on primary targets lst-1 and sygl-1 activity (4 hours auxin treatment). 94 genes are identified to be activated by LAG-1 (48 hours auxin treatment) from 4 biological replicates with the following criteria: gene counts per million greater than 2.0; fold of change greater than 2.0; false discovery rate (FDR) <0.05. Conclusions: lst-1 and sygl-1, two previously reported genes, are identified as primary transcriptional target for LAG-1 in C. elegans germline stem cells. Overall design: Ribo-depleted RNA-seq libraries are prepared with four biological replicates