Use of laser capture microdissection for analysis of retinal mRNA/miRNA expression and DNA methylation

Laszlo Hackler, Tomohiro Masuda, Verity F. Oliver, Shannath L. Merbs, Donald J. Zack

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Laser capture microdissection (LCM) is a useful method to isolate specific cells or cell layers of interest from heterogeneous tissues, such as the retina. The collected cells can be used for DNA, RNA, or protein analysis. We have applied LCM technology to isolate cells from the outer nuclear, inner nuclear, and ganglion cell layers of the retina for mRNA and microRNA (miRNA) expression and epigenetic (DNA methylation) analysis. Here, we describe the methods we have employed for sample preparation, LCM-based isolation of retinal layers, RNA/DNA extraction, RNA quality check, microRNA analysis by quantitative PCR, and DNA methylation analysis by bisulfite sequencing.

Original languageEnglish (US)
Title of host publicationRetinal Development
Subtitle of host publicationMethods and Protocols
PublisherHumana Press Inc.
Pages289-304
Number of pages16
ISBN (Print)9781617798474
DOIs
StatePublished - 2012

Publication series

NameMethods in Molecular Biology
Volume884
ISSN (Print)1064-3745

Keywords

  • Bisulfate sequencing
  • DNA methylation
  • Epigenetics
  • Gene expression
  • Laser capture microdissection
  • Quantitative PCR
  • RNA extraction
  • Retina
  • microRNA

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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  • Cite this

    Hackler, L., Masuda, T., Oliver, V. F., Merbs, S. L., & Zack, D. J. (2012). Use of laser capture microdissection for analysis of retinal mRNA/miRNA expression and DNA methylation. In Retinal Development: Methods and Protocols (pp. 289-304). (Methods in Molecular Biology; Vol. 884). Humana Press Inc.. https://doi.org/10.1007/978-1-61779-848-1_21