Identification of a molecular marker for type A spermatogonia by microarray analysis using gonadal cells from pvasa-GFP transgenic rainbow trout (Oncorhynchus mykiss).
The spermatogonia of fish can be classified as being either undifferentiated type A spermatogonia or differentiated type B spermatogonia. Although type A spermatogonia, which contain spermatogonial stem cells, have been demonstrated to be a suitable material for germ cell transplantation, no molecular markers for distinguishing between type A and type B spermatogonia in fish have been developed to date. We therefore sought to develop a molecular marker for type A spermatogonia in rainbow trout. Using GFP-dependent flow cytometry (FCM), enriched fractions of type A and type B spermatogonia, testicular somatic cells, and primordial germ cells were prepared from rainbow trout possessing the green fluorescent protein (GFP) gene driven by trout vasa regulatory regions (pvasa-GFP rainbow trout). The gene-expression profiles of each cell fraction were then compared with a microarray containing cDNAs representing 16,006 genes from several salmonid species. Genes exhibiting high expression for type A spermatogonia relative to above-mentioned other types of gonadal cells were identified and subjected to RT-PCR and quatitative PCR analysis. Since only the rainbow trout notch1 homologue showed significantly high expression in the type A spermatogonia-enriched fraction, we propose that notch1 may be a useful molecular marker for type A spermatogonia. The combination of GFP-dependent FCM and microarray analysis of pvasa-GFP rainbow trout can therefore be applied to the identification of potentially useful molecular markers of germ cells in fish.
Yano
Ayaka
A
Department of Marine Biosciences, Tokyo University of Marine Science and Technology, 4-5-7 Konan, Minato-ku, Tokyo 108-8477, Japan.
von Schalburg
Kris
K
Cooper
Glenn
G
Koop
Ben F
BF
Yoshizaki
Goro
G
D000818:Animals; D030801:Animals, Genetically Modified; D015415:Biomarkers / Q000378:metabolism*; D020224:Expressed Sequence Tags / Q000378:metabolism; D029941:Fish Proteins / Q000235:genetics / Q000378:metabolism; D005434:Flow Cytometry; D049452:Green Fluorescent Proteins / Q000378:metabolism; D017404:In Situ Hybridization, Fluorescence; D049408:Luminescent Agents / Q000378:metabolism; D008297:Male; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D017686:Oncorhynchus mykiss / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D051881:Receptor, Notch1 / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013093:Spermatogonia / Q000378:metabolism*; D013737:Testis / Q000166:cytology
D015415:Biomarkers; D029941:Fish Proteins; D049408:Luminescent Agents; D051881:Receptor, Notch1; D049452:Green Fluorescent Proteins