have developed a novel, generally-applicable means to analyze cell
type-specific gene expression within eukaryotic organisms (1,2,3). This
involves transgenic expression of GFP (or other Fluorescent Proteins),
targeted to the nucleus and placed under the transcriptional control of
cell type-specific promoters. The nuclei are then released from the
organism by gentle homogenization, are stained with DAPI and are purified
by fluorescence-activated sorting. Expression profiling is then done
using microarrays. We have demonstrated that nuclear transcript levels
are largely concordant with those of total cellular transcripts, both in
plants and animals. In terms of cell type specificity, our results are
consistent with those found through sorting of protoplasts (4,5).
Analyzing and sorting nuclei is of obvious advantage for
species, organs or cells that do not readily produce protoplasts.
In order to expand this technology to
the scientific community at large, we have available the DNA constructions
required for expression and nuclear targeting of GFP. We also are willing
to provide, on a cost-recovery basis, the processes of flow sorting and
nuclear expression profiling. We envisage being sent transgenic seed, which
would be germinated, the plants homogenized, the nuclei purified by
fluorescence-activated sorting, and the nuclear transcripts subjected to
profiling using microarrays or GeneChips following the required
single-round of in vitro amplification. Alternatively, the amplified
RNA would be shipped back to the originating scientist for further
For further information about this
service, and to request the DNA constructions, please contact me by email
or telephone (520.621.9153).
- Galbraith DW (2003). Global analysis of cell
type-specific gene expression. Comparative and Functional
- Zhang CQ, Gong FC, Lambert GM, Galbraith DW (2005).
Cell type-specific characterization of nuclear DNA contents within
complex tissues and organs. Plant Methods 1:7
- Zhang CQ, Lambert GM, Barthelson RA, Galbraith DW
(2008). Characterization of cell-specific gene expression through
fluorescence-activated sorting of nuclei. Plant Physiology 147:30-40.
- Birnbaum K, Shasha DE, Wang JY, Jung JW, Lambert
GM, Galbraith DW, Benfey PN (2003). A gene expression map of the
Arabidopsis root. Science 302:1956-1960.
- Brady SM, Orlando DA, Lee J-Y, Wang JY, Kock J,
Dinneny JR, Mace D, Ohler U, Benfey PN (2007) A high-resolution root
spatiotemporal map reveals dominant expression patterns. Science
Questions or comments should be
addressed to: David Galbraith
Page last updated July 1, 2008