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Related Experiment Video

Updated: Aug 7, 2025

Coculture of Axotomized Rat Retinal Ganglion Neurons with Olfactory Ensheathing Glia, as an In Vitro Model of Adult Axonal Regeneration
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LCM-Seq for Retinal Cell Layer-Specific Responses During Optic Nerve Regeneration.

Wesley Speer1, Matthew B Veldman2

  • 1Department of Cell Biology, Neurobiology, and Anatomy, Medical College of Wisconsin, Milwaukee, WI, USA.

Methods in Molecular Biology (Clifton, N.J.)
|March 7, 2023
PubMed
Summary

This study presents a laser capture microdissection (LCM) method for analyzing gene expression in zebrafish retinal ganglion cells (RGCs) after optic nerve injury. This technique aids in understanding optic nerve regeneration.

Keywords:
Inner nuclear layerLCM-seqMicrodissectionOptic nerveOuter nuclear layerRNA-seqRegenerationRetinaRetinal ganglion cell layerZebrafish

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Area of Science:

  • Neuroscience
  • Molecular Biology
  • Genetics

Background:

  • Retinal ganglion cells (RGCs) are crucial for visual information transmission.
  • Optic nerve injury impairs vision, and regeneration is limited in mammals.
  • Zebrafish offer a model for studying optic nerve regeneration.

Purpose of the Study:

  • To establish a method for laser capture microdissection (LCM) from zebrafish retinal layers.
  • To analyze gene expression changes following optic nerve injury and regeneration.
  • To identify molecular mechanisms underlying successful optic nerve regeneration.

Main Methods:

  • Laser capture microdissection (LCM) of specific retinal layers in zebrafish.
  • RNA purification from microdissected cells.
  • RNA sequencing (RNA-seq) and downstream gene expression analysis.

Main Results:

  • Successful isolation of RNA from distinct retinal layers using LCM.
  • Transcriptome-wide gene expression profiling is achievable with the purified RNA.
  • The method allows for the study of molecular events during optic nerve regeneration.

Conclusions:

  • LCM is a viable technique for studying gene expression in RGCs post-injury.
  • This method facilitates the investigation of molecular pathways in zebrafish optic nerve regeneration.
  • The protocol provides high-quality RNA suitable for various downstream analyses.