RNA profiling of FAC-sorted neurons from the developing zebrafish spinal cord
- Cerda, G.A., Hargrave, M., and Lewis, K.E.
- Developmental dynamics : an official publication of the American Association of Anatomists 238(1): 150-161 (Journal)
- Registered Authors
- Cerda, Gustavo, Hargrave, Murray, Lewis, Katharine E.
- Tg(pax2a:GFP), Tg(elavl3:EGFP), CiA, V1, interneuron, FACS, zebrafish, Affymetrix, RNA-profile, GFP, Lhx1, Lhx5, Eng1b, HuC, Pax2, Elav13
- MeSH Terms
- Animals, Genetically Modified
- Cell Separation*/instrumentation
- Cell Separation*/methods
- Gene Expression Profiling/instrumentation
- Gene Expression Profiling/methods*
- In Situ Hybridization
- Microarray Analysis*/instrumentation
- Microarray Analysis*/methods
- Recombinant Fusion Proteins/genetics
- Recombinant Fusion Proteins/metabolism
- Spinal Cord*/cytology
- Spinal Cord*/embryology
- Spinal Cord*/growth & development
- Zebrafish*/anatomy & histology
- Zebrafish Proteins/genetics
- Zebrafish Proteins/metabolism
- 19097188 Full text @ Dev. Dyn.
Cerda, G.A., Hargrave, M., and Lewis, K.E. (2009) RNA profiling of FAC-sorted neurons from the developing zebrafish spinal cord. Developmental dynamics : an official publication of the American Association of Anatomists. 238(1):150-161.
In this report, we describe a successful protocol for isolating and expression-profiling live fluorescent-protein-labelled neurons from zebrafish embryos. As a proof-of-principle for this method, we FAC-sorted and RNA-profiled GFP-labelled spinal CiA interneurons and compared the expression profile of these cells to those of post-mitotic spinal neurons in general and to all trunk cells. We show that RNA of sufficient quality and quantity to uncover both expected and novel transcription profiles via Affymetrix microarray analysis can be extracted from 5,700 to 20,000 FAC-sorted cells. As part of this study, we also further confirm the genetic homology of mammalian and zebrafish V1 interneurons, by demonstrating that zebrafish V1 cells (CiAs) express genes that encode for the transcription factors Lhx1a and Lhx5. This protocol for dissociating, sorting and RNA-profiling neurons from organogenesis-stage zebrafish embryos should also be applicable to other developing organs and tissues and potentially other model organisms.
Genes / Markers
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Engineered Foreign Genes