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¹H NMR: Interpreting Distorted and Overlapping Signals

¹H NMR: Interpreting Distorted and Overlapping Signals

Spin systems where the difference in chemical shifts of the coupled nuclei is greater than ten times J are called first-order spin systems. These nuclei are weakly coupled, and their chemical shifts and coupling constant can generally be estimated from the well-separated signals in the spectrum.
As Δν decreases and the signals move closer, the doublets appear increasingly distorted. The intensities of the inner lines increase at the cost of those of the outer lines as the signals are...

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Deciphering High-Resolution 3D Chromatin Organization via Capture Hi-C

Deciphering High-Resolution 3D Chromatin Organization via Capture Hi-C

Published on: October 14, 2022

Spectral imaging to visualize higher-order genomic organization.

Iain A Sawyer^1,2, Sergei P Shevtsov¹, Miroslav Dundr¹

¹a Department of Cell Biology , Rosalind Franklin University of Medicine & Science, Chicago Medical School , North Chicago , IL , USA.

Nucleus (Austin, Tex.)

|May 12, 2016

Summary

This study introduces a novel multiplexed DNA FISH technique for analyzing gene pairing and nuclear organization. This method allows simultaneous visualization of multiple genomic structures, advancing high-throughput sequencing data interpretation.

Keywords:

Cajal bodies DNA FISH chromosome territories gene positioning genome organization nuclear bodies spectral imaging spliceosomal U snRNA genes

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Last Updated: Mar 21, 2026

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Published on: October 28, 2025

Area of Science:

Genomics
Cell Biology
Molecular Biology

Background:

Interpreting large, high-throughput sequencing datasets is a key challenge in genomics.
DNA FISH and high-content microscopy validate gene pairing events but have limitations in studying multiple loci simultaneously or their precise nuclear positioning.
Advanced methods are needed to study higher-order nuclear and cellular organization.

Purpose of the Study:

To develop and present a multiplexed DNA FISH technique combined with indirect immunofluorescence.
To enable the study of the relative positions of up to 6 distinct genomic or cellular structures simultaneously.
To validate 3C-based technologies and deepen the understanding of spatial organization within the nucleus.

Main Methods:

A multiplexed DNA FISH technique was developed, integrating indirect immunofluorescence.
Spectral imaging during image acquisition and linear unmixing were employed for simultaneous analysis.
The method was used to quantify gene pairing between highly expressed spliceosomal genes.

Main Results:

The technique allows for the simultaneous study of the relative positions of 6 distinct genomic or cellular structures in a single hybridization step.
Gene pairing between highly expressed spliceosomal genes was quantified and compared to in silico simulations.
This approach offers a universally applicable method for validating other techniques and imaging nuclear organization.

Conclusions:

The developed multiplexed DNA FISH technique overcomes limitations of existing methods for studying nuclear organization.
It provides a powerful tool for validating 3C-based technologies and analyzing spatial relationships of multiple genomic loci.
This method significantly advances the deep imaging of spatial organization within the nucleus and global cellular organization.