Screening for mammalian neural genes via fluorescence-activated cell sorter purification of neural precursors from Sox1-gfp knock-in mice

doi:10.1073/pnas.1734197100

Screening for mammalian neural genes via fluorescence-activated cell sorter purification of neural precursors from Sox1-gfp knock-in mice

^*Institute for Stem Cell Research, University of Edinburgh, King's Buildings, West Mains Road, EH9 3JQ Edinburgh, Scotland; ^‡The Scottish Centre for Genomic Technology and Informatics, University of Edinburgh, Chancellor's Building, 49 Little France Crescent, EH16 4SB Edinburgh, Scotland; and ^§Division of Biomedical Sciences, University of Edinburgh, Hugh Robson Building, George Square, EH8 9XD Edinburgh, Scotland

Abstract

The transcription factor Sox1 is the earliest and most specific known marker for mammalian neural progenitors. During fetal development, Sox1 is expressed by proliferating progenitor cells throughout the central nervous system and in no tissue but the lens. We generated a reporter mouse line in which egfp is inserted into the Sox1 locus. Sox1 ^GFP animals faithfully recapitulate the expression of the endogenous gene. We have used the GFP reporter to purify neuroepithelial cells by fluorescence-activated cell sorting from embryonic day 10.5 embryos. RNAs prepared from Sox1^GFP+ and Sox1^GFP- embryo cells were then used to perform a pilot screen of subtracted cDNAs prepared from differentiating embryonic stem cells and arrayed on a glass chip. Fifteen unique differentially expressed genes were identified, all previously associated with fetal or adult neural tissue. Whole mount in situ hybridization against two genes of previously unknown embryonic expression, Lrrn1 and Musashi2, confirmed the selectivity of this screen for early neuroectodermal markers.

Footnotes

↵ ∥ To whom correspondence should be addressed. E-mail: austin.smith{at}ed.ac.uk.
↵ † J.A. and M.P.S. contributed equally to this work.
↵ ¶ A.S. is a scientific adviser to Stem Cell Sciences, Ltd., and holds nonvoting equity in the company. Stem Cell Sciences funds research in the laboratory and has patents granted and pending on technology used in this article.
This paper results from the Arthur M. Sackler Colloquium of the National Academy of Sciences, “Regenerative Medicine,” held October 18-22, 2002, at the Arnold and Mabel Beckman Center of the National Academies of Science and Engineering in Irvine, CA.
Abbreviations: ES, embryonic stem; FACS, fluorescence-activated cell sorting; En, embryonic day n.