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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Reference Gene Selection for Gene Expression Analysis of Oocytes Collected from Dairy Cattle and Buffaloes during Winter and Summer

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Macabelli, Carolina Habermann [1, 2] ; Ferreira, Roberta Machado [2, 3] ; Gimenes, Lindsay Unno [4] ; Tonizza de Carvalho, Nelcio Antonio [5] ; Soares, Julia Gleyci [3] ; Ayres, Henderson [3, 6] ; Ferraz, Marcio Leao [7] ; Watanabe, Yeda Fumie [8] ; Watanabe, Osnir Yoshime [8] ; Sangalli, Juliano Rodrigues [2, 9] ; Smith, Lawrence Charles [10] ; Baruselli, Pietro Sampaio [3] ; Meirelles, Flavio Vieira [2, 9] ; Chiaratti, Marcos Roberto [2, 9, 1]
Total Authors: 14
Affiliation:
[1] Univ Fed Sao Carlos, Ctr Ciencias Biol & Saude, Dept Genet & Evolucao, BR-13560 Sao Carlos, SP - Brazil
[2] Univ Sao Paulo, Fac Zootecnia & Engn Alimentos, Dept Med Vet, Pirassununga, SP - Brazil
[3] Univ Sao Paulo, Fac Med Vet & Zootecnia, Dept Reprod Anim, Sao Paulo - Brazil
[4] Univ Estadual Paulista, Fac Ciencias Agr & Vet, Dept Reprod Anim, Jaboticabal, SP - Brazil
[5] APTA, Unidade Pesquisa Desenvolvimento Registro, Registro, SP - Brazil
[6] Vida Reprod Consultoria, Cravinhos, SP - Brazil
[7] MSD Saude Anim, Sao Paulo - Brazil
[8] LTDA, WTA Tecnol Aplicada, Cravinhos, SP - Brazil
[9] Univ Sao Paulo, Fac Med Vet & Zootecnia, Dept Cirurgia, Sao Paulo - Brazil
[10] Univ Montreal, Fac Med Vet, Ctr Rech Reprod Anim, St Hyacinthe, PQ J2S 7C6 - Canada
Total Affiliations: 10
Document type: Journal article
Source: PLoS One; v. 9, n. 3 MAR 27 2014.
Web of Science Citations: 13
Abstract

Oocytes from dairy cattle and buffaloes have severely compromised developmental competence during summer. While analysis of gene expression is a powerful technique for understanding the factors affecting developmental hindrance in oocytes, analysis by real-time reverse transcription PCR (RT-PCR) relies on the correct normalization by reference genes showing stable expression. Furthermore, several studies have found that genes commonly used as reference standards do not behave as expected depending on cell type and experimental design. Hence, it is recommended to evaluate expression stability of candidate reference genes for a specific experimental condition before employing them as internal controls. In acknowledgment of the importance of seasonal effects on oocyte gene expression, the aim of this study was to evaluate the stability of expression levels of ten well-known reference genes (ACTB, GAPDH, GUSB, HIST1H2AG, HPRT1, PPIA, RPL15, SDHA, TBP and YWHAZ) using oocytes collected from different categories of dairy cattle and buffaloes during winter and summer. A normalization factor was provided for cattle (RPL15, PPIA and GUSB) and buffaloes (YWHAZ, GUSB and GAPDH) based on the expression of the three most stable reference genes in each species. Normalization of non-reference target genes by these reference genes was shown to be considerably different from normalization by less stable reference genes, further highlighting the need for careful selection of internal controls. Therefore, due to the high variability of reference genes among experimental groups, we conclude that data normalized by internal controls can be misleading and should be compared to not normalized data or to data normalized by an external control in order to better interpret the biological relevance of gene expression analysis. (AU)

FAPESP's process: 11/14207-0 - Effect of season on oocyte quality of buffaloes (Bubalus bubalis): analysis of gene expression and mitochondrial DNA copy number
Grantee:Carolina Habermann Macabelli
Support type: Scholarships in Brazil - Scientific Initiation
FAPESP's process: 10/09561-7 - Effect of introducing a large amount of embryonic or somatic mitochondria on development and on mitochondrial inheritance in bovines
Grantee:Marcos Roberto Chiaratti
Support type: Scholarships in Brazil - Post-Doctorate
FAPESP's process: 09/00938-3 - Effect of season on oocyte quality and in vitro embryo production in Holstein cows (Bos taurus) of different categories
Grantee:Pietro Sampaio Baruselli
Support type: Regular Research Grants
FAPESP's process: 12/07510-1 - Molecular evaluation of oocyte maturation and use of cytoplasmic transfer to increse oocyte competence of repeat breeder dairy cows
Grantee:Roberta Machado Ferreira Saran
Support type: Scholarships in Brazil - Post-Doctorate
FAPESP's process: 10/13384-3 - Effect of introducing embryonic or somatic mitochondria on development and on mitochondrial inheritance in cattle
Grantee:Flávio Vieira Meirelles
Support type: Regular Research Grants