An mRNA differential display strategy for cloning genes expressed during mouse gonad development.
The mRNA differential display technique has become a popular method for isolating novel genes in a variety of biological systems including carcinogenesis, hormone regulation, plant biology and neurobiology. We have further developed the method by optimizing different steps for the use of small amounts of material, such that differential display can be used in the study of developmental biology. Our techniques include a new assay for elimination of false positive cDNA clones and a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) method for the rapid analysis of differences in gene expression. This improved mRNA differential display strategy requires less than 4 microg of total RNA. We have used it for the isolation of genes which are expressed during gonad development in the mouse. One of the cDNAs found, cDNA 4.3 which corresponds to a part of the gene encoding the steroid hydroxylase 3betaHSD I, was shown to be a valuable marker for adrenal development and for Leydig cell differentiation and organization during testis development.
Nordqvist
K
K
Department of Cell and Molecular Biology, Medical Nobel Institute, Karolinska Institutet, Stockholm, Sweden. Katarina.Nordqvist@cmb.ki.se
Töhönen
V
V
D015096:3-Hydroxysteroid Dehydrogenases / Q000235:genetics; D000311:Adrenal Glands / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D000818:Animals; D002454:Cell Differentiation; D003001:Cloning, Molecular / Q000379:methods*; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000032:analysis / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D018507:Gene Expression Regulation, Developmental; D006066:Gonads / Q000166:cytology / Q000196:embryology* / Q000378:metabolism; D017403:In Situ Hybridization; D007985:Leydig Cells / Q000166:cytology / Q000378:metabolism; D008297:Male; D051379:Mice; D009687:Nuclear Proteins; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D017422:Sequence Analysis, DNA; D051876:Sex-Determining Region Y Protein; D013254:Steroid 17-alpha-Hydroxylase / Q000235:genetics; D013737:Testis / Q000196:embryology / Q000378:metabolism; D014157:Transcription Factors
D017931:DNA Primers; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D051876:Sex-Determining Region Y Protein; D014157:Transcription Factors; D015096:3-Hydroxysteroid Dehydrogenases; D013254:Steroid 17-alpha-Hydroxylase