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Related Experiment Video

Updated: Sep 8, 2025

Simultaneous Mapping and Quantitation of Ribonucleotides in Human Mitochondrial DNA
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Targeted Nuclease Approaches for Mapping Native R-Loops.

Phillip Wulfridge1, Qingqing Yan1, Kavitha Sarma2

  • 1Gene Expression and Regulation Program, The Wistar Institute, Philadelphia, PA, USA.

Methods in Molecular Biology (Clifton, N.J.)
|June 15, 2022
PubMed
Summary

MapR and BisMapR are novel methods for detecting DNA:RNA hybrid R-loops genome-wide. These techniques accurately map R-loop locations and strand of origin, aiding gene regulation and disease research.

Keywords:
DNA:RNA hybridDeep sequencingR-loopStrand-specific

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Area of Science:

  • Molecular Biology
  • Genomics
  • Epigenetics

Background:

  • R-loops, which are DNA:RNA hybrid structures, form naturally during transcription.
  • Understanding R-loop genomic locations and origin is crucial for studying gene regulation and disease.
  • Existing methods for R-loop detection have limitations in resolution, specificity, or input requirements.

Purpose of the Study:

  • To develop a novel, high-resolution, genome-wide method for detecting R-loops.
  • To enable strand-specific identification of R-loop locations.
  • To provide a robust protocol with low cell input and short experimental time.

Main Methods:

  • Development of MapR, a nuclease-based protocol using a modified RNase H enzyme for R-loop cleavage and deep sequencing.
  • Extension of MapR to BisMapR, incorporating non-denaturing bisulfite conversion for enhanced strand specificity.
  • Application of these methods for genome-wide R-loop profiling.

Main Results:

  • MapR and BisMapR successfully identify R-loops with high resolution and low background noise.
  • The methods are effective with low cellular input and require minimal experimental time.
  • Strand-specific R-loop mapping is achieved, providing precise genomic localization.

Conclusions:

  • MapR and BisMapR offer significant advancements in R-loop detection technology.
  • These methods facilitate a deeper understanding of R-loop functions in biological processes.
  • The developed protocols are valuable tools for research in gene regulation and associated diseases.