title: Reference genes to quantify gene expression during oogenesis in a teleost fish.
abstract: Understanding the molecular events involved in the acquisition of competence during oogenesis is a key step to determine the secret of 'high quality' eggs for aquaculture. Quantitative real time polymerase chain reaction (qPCR) is the technique of election to determine changes in transcript abundance in such studies, but choosing reference genes for normalization, in particular during oogenesis, remains a challenge. In the present study, transcription of 6 functionally distinct genes, β actin (ACTB), cathepsin D (CTSD), cathepsin Z (CTSZ), elongation factor 1 α (EEF1A), TATA binding protein (TBP) and tubulin A (TUBA1A) was assessed as normalizers of bone morphogenetic protein (BMP) and activin membrane-bound inhibitor (BAMBI) gene expression in mRNA from Mozambique tilapia oocytes during oogenesis. Reverse transcription was equally efficient and varies little in all samples. Most of the genes considered for reference were stable during early stages of oogenesis but variations were observed during vitellogenesis. A single gene and up to 3 genes were shown to be insufficient for reliable normalization throughout the whole oogenesis. The combination of the genes ACTB, CTSD, EEF1A and CTSZ as reference was found to minimize variation and has the most stable expression pattern between maturation stages.
authors: Centre of Marine Sciences (CCMar), University of the Algarve, Campus de Gambelas, 8005-139 Faro, Portugal. deloffre@ualg.pt
authors: Laurence A M
authors: LA
authors: Deloffre
authors: André
authors: A
authors: Andrade
authors: Alexandra I
authors: AI
authors: Filipe
authors: Adelino V M
authors: AV
authors: Canario
mesh_terms: D000199:Actins / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D002402:Cathepsin D / Q000235:genetics; D056663:Cathepsin Z / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D005260:Female; D029941:Fish Proteins / Q000235:genetics; D015870:Gene Expression; D005819:Genetic Markers; D009866:Oogenesis / Q000235:genetics*; D020648:Peptide Elongation Factor 1 / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D035181:TATA-Box Binding Protein / Q000235:genetics; D017210:Tilapia / Q000235:genetics* / Q000502:physiology; D014404:Tubulin / Q000235:genetics
chemical_list: D000199:Actins; D017931:DNA Primers; D029941:Fish Proteins; D005819:Genetic Markers; D020648:Peptide Elongation Factor 1; D035181:TATA-Box Binding Protein; D014404:Tubulin; D056663:Cathepsin Z; D002402:Cathepsin D