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Validation of reference genes for ovarian tissue from capuchin monkeys (Cebus apella)

Published online by Cambridge University Press:  05 April 2012

A.B. Brito ,
J.S. Lima ,
D.C. Brito ,
L.N. Santana ,
N.N. Costa ,
M.S. Miranda ,
O.M. Ohashi ,
R.R. Santos  and
S.F.S. Domingues
A.B. Brito
Affiliation:
Laboratory of Wild Animal Biology and Medicine, Universidade Federal do Pará, Brazil. Animal Science Post-graduation Program, Universidade Federal do Pará, Brazil.
J.S. Lima
Affiliation:
Laboratory of Wild Animal Biology and Medicine, Universidade Federal do Pará, Brazil. Animal Science Post-graduation Program, Universidade Federal do Pará, Brazil.
D.C. Brito
Affiliation:
Laboratory of Wild Animal Biology and Medicine, Universidade Federal do Pará, Brazil.
L.N. Santana
Affiliation:
Laboratory of Wild Animal Biology and Medicine, Universidade Federal do Pará, Brazil. Animal Science Post-graduation Program, Universidade Federal do Pará, Brazil.
N.N. Costa
Affiliation:
Institute of Biological Sciences, Universidade Federal do Pará, Brazil.
M.S. Miranda
Affiliation:
Institute of Biological Sciences, Universidade Federal do Pará, Brazil.
O.M. Ohashi
Affiliation:
Institute of Biological Sciences, Universidade Federal do Pará, Brazil.
R.R. Santos*
Affiliation:
Rua Augusto Corrêa, Campus Básico, CEP 66075–110, Belém, Pará, Brazil. Laboratory of Wild Animal Biology and Medicine, Universidade Federal do Pará, Brazil. Animal Science Post-graduation Program, Universidade Federal do Pará, Brazil. Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.
S.F.S. Domingues
Affiliation:
Laboratory of Wild Animal Biology and Medicine, Universidade Federal do Pará, Brazil. Animal Science Post-graduation Program, Universidade Federal do Pará, Brazil.
*
All correspondence to: Regiane R dos Santos. Rua Augusto Corrêa, Campus Básico, CEP 66075–110, Belém, Pará, Brazil. Tel:/Fax: +55 81 32018011. e-mail: [email protected]
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Summary

There is no tradition in studies reporting the effect of exposure to cryoprotectants or simply hypoxia and hypothermia on gene expression in the ovarian tissue and there has been only one study on reference or target genes quantification, and comparisons of normoxic with hypoxic, hypothermic and toxic conditions. Our aim in the present study was to investigate the stability of three reference genes in the ovarian tissue of capuchin monkeys (Cebus apella). To this end, fresh and cryoprotectant-exposed ovarian biopsies were used. Both fresh and exposed ovarian tissues were subjected to total RNA extraction and synthesis of cDNA. cDNA was amplified by real-time polymerase chain reaction (PCR), and GeNorm, BestKeeper and NormFinder software were used to evaluate the stability of glyceraldehyde-2-phosphate dehydrogenase (GAPDH), hypoxanthine phosphoribosyltransferase 1 (HPRT1) and TATA-binding protein (TBP). Results demonstrated that, in the ovarian tissue from capuchin monkeys, HPRT1 and TBP were the most suitable reference genes and thus could be used as parameters to normalize data in future studies. In contrast, GAPDH appeared as the least stable gene among the tested reference genes. In conclusion, HPRT1 and TBP were the most stable reference genes in fresh and cryoprotectant-exposed ovarian tissue from capuchin monkeys.

Keywords

GAPDHHPRT1Ovarian tissueqRT-PCRStably expressed genesTBP
Type
Research Article
Information
Zygote , Volume 21 , Issue 2 , May 2013 , pp. 167 - 171
Copyright
Copyright © Cambridge University Press 2012

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