GAPDH pseudogenes and the quantification of feline genomic DNA equivalents
| Autor: | Stefan Widmer, Regina Hofmann-Lehmann, A. Katrin Helfer-Hungerbuehler |
|---|---|
| Přispěvatelé: | University of Zurich |
| Jazyk: | angličtina |
| Rok vydání: | 2013 |
| Předmět: |
Genetics
0303 health sciences Article Subject biology 630 Agriculture 040301 veterinary sciences Pseudogene Cell 04 agricultural and veterinary sciences Genome 0403 veterinary science 10187 Department of Farm Animals 03 medical and health sciences genomic DNA Equivalent medicine.anatomical_structure stomatognathic system Single copy gene biology.protein medicine Nucleic acid 570 Life sciences Glyceraldehyde 3-phosphate dehydrogenase 030304 developmental biology Research Article |
| Zdroj: | Molecular Biology International |
| Popis: | Quantitative real-time PCR (qPCR) is broadly used to detect and quantify nucleic acid targets. In order to determine cell copy number and genome equivalents, a suitable reference gene that is present in a defined number in the genome is needed, preferably as a single copy gene. For most organisms, a variable number of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) pseudogenes have been reported. However, it has been suggested that a single-copy of the GAPDH pseudogene is present in the feline genome and that a GAPDH assay can therefore be used to quantify feline genomic DNA (gDNA). The aim of this study was to determine whether one or more GAPDH pseudogenes are present in the feline genome and to provide a suitable alternative qPCR system for the quantification of feline cell copy number and genome equivalents. Bioinformatics and sequencing results revealed that not just one but several closely related GAPDH-like sequences were present in the cat genome. We thus identified, developed, optimized, and validated an alternative reference gene assay using feline albumin (fALB). Our data emphasize the need for an alternative reference gene, apart from the GAPDH pseudogene, for the normalization of gDNA levels. We recommend using the fALB qPCR assay for future studies. |
| Databáze: | OpenAIRE |
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