Project description:Stroma extracts were isolated from 2-week-old transgenic dPPRrbcL or WT plants and incubated with HA-specific antibodies. IgGs were captured with Protein A Dynabeads (Thermo Fisher scientific) and recovered RNA was used for generation of libraries with the NEBNext® Ultra™ II RNA Library Prep Kit. Primary reads were aligned to the Arabidopsis chloroplast genome (accession number NC_000932.1) using CLC Genomics Workbench. Reads of the two replicates aligned in CLC Genomics Workbench were extracted as coverage (reads per nucleotide). Graphs were created with excel using the extracted coverage values
Project description:blanc09_ripseq_rfl8-ripseq experiment for rfl8 targets-Which mitochondrial transcripts are bound by RFL8 protein? -The RIPseq approach to find the mitochondrial transcripts attached by RFL8 protein via co-immunoprecipitation of 3HA tag fused in C-terminal of RFL8 protein
Project description:Stroma extracts were isolated from 2-week-old WT plants and incubated with either PUMPKIN-specific antibodies or with the pre-immune serum. IgGs were captured with SiMAG-Protein G beads (Chemicell) and recovered RNA was used for generation of libraries with the ScriptSeq v2 RNA-seq Library Preparation Kit (Epicentre). Primary reads were aligned to the Arabidopsis chloroplast genome (accession number NC_000932.1) using CLC Genomics Workbench, the mean RPKM values of the replicates as well as the ratio of PUMPKIN vs Pre-immuneserum were calculated. Five prominent RNA targets (trnG-UCC, trnV-UAC, petB, petD and ndhA) were identified and validated via Slot Blot analyses.
Project description:Stroma extracts were isolated from 2-week-old WT plants and incubated with either BSF-specific antibodies or with the pre-immune serum. IgGs were captured with SiMAG-Protein G beads (Chemicell) and recovered RNA was used for generation of libraries with the ScriptSeq v2 RNA-seq Library Preparation Kit (Epicentre). Primary reads were aligned to the Arabidopsis chloroplast genome (accession number NC_000932.1) using CLC Genomics Workbench. BAM files were extracted and sorted in Galaxy . Sorted BAM files were converted into RPKM-normalized bigwig files and displayed in IGB. The differential enrichment of BSF/control of the two replicates was displayed across the entire chloroplast genome to identify the RNA targets of BSF.