A systematic molecular genetic approach to study mammalian germline development

Kuniya Abe, Minoru Ko, Grant R. Macgregor

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

It is difficult to study gene expression in mammalian embryonic germ cells as PGCs constitute only a minor proportion of the mouse embryo. We have overcome this problem by using a novel combination of established molecular and transgenic approaches. A line of mice has been generated in which the cells of the germ lineage express the β-galactosidase reporter gene during embryogenesis. Using this line, germ cells have been purified to near homogeneity from embryos at discrete stages during germline development by use of a stain for β-gal activity and a fluorescence activated cell sorter. Subsequently, cDNA libraries have been constructed from each germ cell population using a modified lone-linker PCR strategy. These combined cDNA libraries represent genes expressed in PGCs during mammalian germline development. To facilitate a molecular genetic approach to studying mammalian germline development, these cDNA libraries will be pooled to form an arrayed, addressed reference embryonic germ cell cDNA library. In parallel with large- scale cDNA sequencing efforts, genes that are differentially expressed in germ cells will be identified by screening the reference library with probes generated by subtractive hybridization. Complementary DNAs identified using this approach will be analyzed by sequencing, database comparison, genomic mapping and in situ hybridization to ascertain the potential functional importance of each gene to germline development. In addition to providing a wealth of novel information regarding patterns of gene expression during mammalian germline development, these results will form the basis for future experiments to determine the function of these genes in this process.

Original languageEnglish
Pages (from-to)1051-1065
Number of pages15
JournalInternational Journal of Developmental Biology
Volume42
Issue number7
Publication statusPublished - 1998
Externally publishedYes

Fingerprint

Gene Library
Molecular Biology
Germ Cells
Genes
Embryonic Structures
Complementary DNA
Galactosidases
Gene Expression
Cell Lineage
Reporter Genes
Libraries
Embryonic Development
In Situ Hybridization
Coloring Agents
Fluorescence
Databases
Polymerase Chain Reaction
Population
Embryonic Germ Cells

Keywords

  • Embryogenesis
  • Germline
  • Mammalian
  • Molecular genetic

ASJC Scopus subject areas

  • Developmental Biology
  • Cell Biology

Cite this

A systematic molecular genetic approach to study mammalian germline development. / Abe, Kuniya; Ko, Minoru; Macgregor, Grant R.

In: International Journal of Developmental Biology, Vol. 42, No. 7, 1998, p. 1051-1065.

Research output: Contribution to journalArticle

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