Simultaneous profiling of chromatin-associated RNA at targeted DNA loci and RNA-RNA Interactions through TaDRIM-seq.

Eukaryotic genomes are extensively transcribed into various types of RNAs, many of which are physically associated with chromatin in cis at their transcription sites or in trans to other genomic loci. Emerging roles have been uncovered for these chromatin-associated RNAs (caRNAs) in gene regulation and genome organization, yet they remain challenging to interrogate. Here, we present TaDRIM-seq, a technique employing Protein G (PG)-Tn5-targeted DNA elements and in situ proximity ligation to concurrently probe caRNAs across diverse genomic regions as well as global RNA-RNA interactions within intact nuclei. Notably, this approach diminishes required cell inputs, minimizes hands-on time compared to established methodologies, and is compatible in both mammalian cells and plants. Using this technique, we identify extensive caRNAs at DNA anchor regions associated with chromatin loops and reveal diurnal variation in RNA-DNA and RNA-RNA connectivity networks within rice.