Current methods to identify sites of RNA interaction with the genome are limited to the study of a single RNA at a time. Here we describe a protocol for ChAR-seq, a strategy to identify all chromatin-associated RNAs and map their DNA contacts genome-wide. In ChAR-seq, proximity ligation ...
For subsequent studies, only the bands resulting from the main PARP1-bound RNA bands (~140 kDa) were used. Seven biological replica experiments were per- formed, barcoded, and pooled for sequencing using PE Illumina sequencing on a HiSeq 2500. From the Manana Melikishvili et al. 5 ...
Processing of PAR-CLIP RNA samples for sequencing. (a) Outline of experiments. PAR-CLIP of endogenous PARP1 was performed on HeLa cells using 4-thiouridine (4SU). (b) PAR-CLIP samples run on an SDS PAGE gel were imaged with Typhoon. (c) Protein-bound samples were transferred onto a ni...
ChAR-seqChromatinNext-generation sequencingNon-coding RNAProximity ligationRNATranscriptomeRNAs play key roles in the cell as molecular intermediates for protein synthesis and as regulators of nuclear processes such as splicing, posttranscriptional regulation, or chromatin remodeling...doi...
4.5. RNA-seq Library Generation, Sequencing, and Mapping Total RNA of GT and leaves were extracted by TRIzol (Invitrogen, Waltham, MA, USA) and mRNA was purified by the Dynabeads mRNA DIRECT kit (Invitrogen). First-strand DNA was synthesized with reverse transcription with oligo dT as a ...