Format

Send to

Choose Destination
Nat Protoc. 2019 Nov;14(11):3243-3272. doi: 10.1038/s41596-019-0229-4. Epub 2019 Oct 16.

Mapping RNA-chromatin interactions by sequencing with iMARGI.

Author information

1
Department of Bioengineering, University of California, San Diego, La Jolla, CA, USA.
2
Institute of Engineering in Medicine, University of California, San Diego, La Jolla, CA, USA.
3
Department of Diabetes Complications and Metabolism, Beckman Research Institute, City of Hope, Duarte, CA, USA.
4
Department of Bioengineering, University of California, San Diego, La Jolla, CA, USA. szhong@ucsd.edu.
5
Institute of Engineering in Medicine, University of California, San Diego, La Jolla, CA, USA. szhong@ucsd.edu.

Abstract

RNA-chromatin interactions represent an important aspect of the transcriptional regulation of genes and transposable elements. However, analyses of chromatin-associated RNAs (caRNAs) are often limited to one caRNA at a time. Here, we describe the iMARGI (in situ mapping of RNA-genome interactome) technique, which is used to discover caRNAs and reveal their respective genomic interaction loci. iMARGI starts with in situ crosslinking and genome fragmentation, followed by converting each proximal RNA-DNA pair into an RNA-linker-DNA chimeric sequence. These chimeric sequences are subsequently converted into a sequencing library suitable for paired-end sequencing. A standardized bioinformatic software package, iMARGI-Docker, is provided to decode the paired-end sequencing data into caRNA-DNA interactions. Compared to its predecessor MARGI (mapping RNA-genome interactions), the number of input cells for iMARGI is 3-5 million (a 100-fold reduction), experimental time is reduced, and clear checkpoints have been established. It takes a few hours a day and a total of 8 d to complete the construction of an iMARGI sequencing library and 1 d to carry out data processing with iMARGI-Docker.

PMID:
31619811
DOI:
10.1038/s41596-019-0229-4

Supplemental Content

Full text links

Icon for Nature Publishing Group
Loading ...
Support Center