Transcription profiling by high throughput sequencing of Arabidopsis LAS-expressing organ boundary cells and AS1-expressing leaf primordia and cotyledon cells

To study cell type-specific gene expression in the leaf boundary region in the genome scale, we employed the TRAP-seq approach recently implemented. In brief, we introduced a reporter line carrying the fusion of the large subunit ribosomal protein L18 with N-terminal His and FLAG epitope tags (HF-RPL18) under the control of the pOp promoter into driver lines expressing the chimeric TF LhG4 under the control of the LATERAL SUPPRESSOR (LAS) promoter, and under the control of the ASYMMETRIC LEAVES1 (AS1) promoter. These driver lines were chosen because pLAS::LhG4 has boundary region-specific activity (Goldshmidt et al, 2008), and pAS1::LhG4 drives pOp reporter expression throughout emerging leaf primordia, but not in the SAM. Cell type-specific expression of HF-RPL18 can efficiently incorporate epitope tags into polysomes for immunopurification of all translating cellular mRNAs. We immunopurified polysomes from seedlings at 7 days after germination (DAG), to isolate translating mRNA in the LAS-expressing organ boundary cells and AS1-expressing leaf primordia and cotyledon cells. Then, we used deep sequencing to map and quantify these mRNA samples.