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Selection of porcine oocytes in vitro through brilliant cresyl blue staining in distinct incubation media

Published online by Cambridge University Press:  13 December 2016

Elisa C.S Santos ,
Jorgea Pradieé ,
Elisângela M. Madeira ,
Miriane M. Pereira ,
Bruna Mion ,
Rafael G. Mondadori ,
Arnaldo D. Vieira ,
Ligia M.C. Pegoraro  and
Thomaz Lucia Jr
Elisa C.S Santos
Affiliation:
ReproPel, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil. Centro de Desenvolvimento Tecnológico, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil.
Jorgea Pradieé
Affiliation:
ReproPel, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil. Faculdade de Veterinária, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil.
Elisângela M. Madeira
Affiliation:
ReproPel, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil. Faculdade de Veterinária, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil.
Miriane M. Pereira
Affiliation:
Embrapa Clima Temperado, 96010–900, Pelotas, RS, Brasil.
Bruna Mion
Affiliation:
ReproPel, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil. Faculdade de Veterinária, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil.
Rafael G. Mondadori
Affiliation:
ReproPel, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil. Instituto de Biologia, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil
Arnaldo D. Vieira
Affiliation:
ReproPel, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil. Faculdade de Veterinária, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil.
Ligia M.C. Pegoraro
Affiliation:
Embrapa Clima Temperado, 96010–900, Pelotas, RS, Brasil.
Thomaz Lucia Jr*
Affiliation:
ReproPel, Faculdade de Veterinária, Universidade Federal de Pelotas, 96010–900-Pelotas-RS, Brazil. Faculdade de Veterinária, Universidade Federal de Pelotas, 96010–900, Pelotas, RS, Brazil.
*
All correspondence to: Thomaz Lucia Jr. ReproPel, Faculdade de Veterinária, Universidade Federal de Pelotas, 96010–900-Pelotas-RS, Brazil. E-mail: thomaz@pq.cnpq.br
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Summary

Staining with brilliant cresyl blue (BCB) may be used for oocyte selection, but BCB staining itself and the most commonly used selection medium (DMPBS) may compromise the development of porcine oocytes in vitro. This study evaluated DNA fragmentation, nuclear maturation, the area of migration of cortical granules (CG) and embryo development for stained (BCB+) and unstained (BCB−) oocytes incubated in DMPBS and in a modified medium (ReproPel) tested for the first time. Unexposed (UN), BCB+ and BCB− oocytes were incubated composing six groups: DMPBS/UN; DMPBS/BCB+; DMPBS/BCB−; ReproPel/UN; ReproPel/BCB+; and ReproPel/BCB−. There were more BCB+ oocytes in ReproPel than in DMPBS (P < 0.05). The DNA fragmentation was evaluated for oocytes in DMPBS/BCB+, DMPBS/BCB−, ReproPel/BCB+, ReproPel/BCB− and in porcine follicular fluid (control). The frequency of oocytes with no DNA fragmentation was greatest (64.6%) in DMPBS/BCB+ and lowest in ReproPel/BCB+ and ReproPel/BCB− (26.8 and 34.1%, respectively) (P < 0.05). Nuclear maturation rates were greater (P < 0.05) for DMPBS/BCB+ (63.1%), ReproPel/UN (55.1%) and ReproPel/BCB+ (50.2%) than for DMPBS/UN (40.8%) and ReproPel/BCB− (35.5%). The area of CG was greater (P < 0.05) for ReproPel/BCB− (80.7%) and DMPBS/UN (77.6%) than for ReproPel/UN (34.7%). Cleavage rates for DMPBS/BCB+ and ReproPel/BCB+ were greater than for DMPBS/UN (P < 0.05). Blastocyst development rates were greatest (P < 0.05) for ReproPel/UN and ReproPel/BCB+. In both media, BCB staining was apparently unable to select competent oocytes, which likely occurred due to toxicity. Despite the similar nuclear maturation and area of CG compared with DMPBS, oocytes selected in ReproPel presented impaired DNA integrity.

Keywords

DNA fragmentationMaturationMediaPorcine oocytesToxicity
Type
Research Article
Information
Zygote , Volume 25 , Issue 1 , February 2017 , pp. 49 - 55
Copyright
Copyright © Cambridge University Press 2016 

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