Research article summary (published 4 Aug 2009):

Obtaining high quality RNA from single cell populations in human postmortem brain tissue.

Full Abstract

We proposed to investigate the gray matter reduction in the superior temporal gyrus seen in schizophrenia patients, by interrogating gene expression profiles of pyramidal neurons in layer III. It is well known that the cerebral cortex is an exceptionally heterogeneous structure comprising diverse regions, layers and cell types, each of which is characterized by distinct cellular and molecular compositions and therefore differential gene expression profiles. To circumvent the confounding effects of tissue heterogeneity, we used laser-capture microdissection (LCM) in order to isolate our specific cell-type i.e pyramidal neurons. Approximately 500 pyramidal neurons stained with the Histogene staining solution were captured using the Arcturus XT LCM system. RNA was then isolated from captured cells and underwent two rounds of T7-based linear amplification using Arcturus/Molecular Devices kits. The Experion LabChip (Bio-Rad) gel and electropherogram indicated good quality a(m)RNA, with a transcript length extending past 600nt required for microarrays. The amount of mRNA obtained averaged 51 microg, with acceptable mean sample purity as indicated by the A260/280 ratio, of 2.5. Gene expression was profiled using the Human X3P GeneChip probe array from Affymetrix.

Author information

Author/s: Pietersen, Charmaine Y (CY); Lim, Maribel P (MP); Woo, Tsung-Ung W (TU);

Affiliation: Department of Structural and Molecular Neuroscience, McLean Hospital. cpietersen(-atsign-)mclean.harvard.edu

Grants: P50MH080272 (Agency:NIMH NIH HHS) ; R01MH76060 (Agency:NIMH NIH HHS)

Journal and publication information

Publication Type: Interactive Tutorial; Research Support, N.I.H., Extramural

Journal: Journal of visualized experiments : JoVE (J Vis Exp), published in United States. (Language: eng)

Reference: 2009-; vol (issue 30) : pp

Dates: Created 2009/08/17; Completed 2009/10/15;

PMID: 19684564, status: MEDLINE (last retrieval date: 10/15/2009, IMS Date: )

Sourced from the National Library of Medicine. Abstract text and other information may be subject to copyright.

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MeSH headings (categories)

This article was linked to the MESH Headings shown below.

Brain - cytology; physiology Brain Chemistry Gene Expression Profiling Humans

Microdissection - methods Oligonucleotide Array Sequence Analysis Pyramidal Cells - chemistry RNA - isolation & purification

Associated Chemicals: RNA (63231-63-0)

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