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Frontiers of Agricultural Science and Engineering

Front. Agr. Sci. Eng.    2019, Vol. 6 Issue (1) : 61-65     https://doi.org/10.15302/J-FASE-2018227
RESEARCH ARTICLE
Effects of enucleation method on in vitro and in vivo development rate of cloned pig embryos
Chengcheng ZHAO1,2, Junsong SHI3, Rong ZHOU3, Ranbiao MAI3, Lvhua LUO3, Xiaoyan HE3, Hongmei JI3, Gengyuan CAI1,2, Dewu LIU1,2, Enqin ZHENG1,2, Zhenfang WU1,2(), Zicong LI2()
1. National Engineering Research Center for Breeding Swine Industry, College of Animal Science, South China Agricultural University, Guangzhou 510642, China
2. Guangdong Provincial Key Laboratory of Agro-Animal Genomics and Molecular Breeding, College of Animal Science, South China Agricultural University, Guangzhou 510642, China
3. Guangdong Wens Foodstuff Group Co., Ltd., Yunfu 527400, China
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Abstract

Enucleation is a crucial procedure for mammalian somatic cell nuclear transfer (SCNT), especially for domestic animal cloning. Oocytes of domestic animals such as pigs and cattle contain dark lipid droplets that hinder localization and removal of the nucleus. Using an oocyte enucleation technique that can obtain a high enucleation rate but has minimal negative effects on the reprogramming potential of oocyte for cloning is beneficial for enhancing the outcome of SCNT. In this study, we compared the pig cloning efficiency resulting from blind aspiration-based (BA-B) enucleation and spindle imaging system-assisted (SIS-A) enucleation, and compared the pig SCNT success rate associated with BA-B enucleation and blind aspiration plus post-enucleation staining-based (BAPPS-B) enucleation. SIS-A enucleation achieved a significantly higher oocyte enucleation success rate and tended to obtain a higher in vivo full term development rate of SCNT embryos than BA-B enucleation. BAPPS-B enucleation also obtained significantly higher in vitro as well as in vivo full term development efficiency of cloned porcine embryos than BA-B enucleation. These data indicate that SIS-A and BAPPS-B enucleation are better approaches for pig SCNT than BA-B enucleation.

Keywords cloning      enucleation      pig      SCNT     
Corresponding Author(s): Zhenfang WU,Zicong LI   
Just Accepted Date: 10 May 2018   Online First Date: 04 June 2018    Issue Date: 25 February 2019
 Cite this article:   
Chengcheng ZHAO,Junsong SHI,Rong ZHOU, et al. Effects of enucleation method on in vitro and in vivo development rate of cloned pig embryos[J]. Front. Agr. Sci. Eng. , 2019, 6(1): 61-65.
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http://journal.hep.com.cn/fase/EN/10.15302/J-FASE-2018227
http://journal.hep.com.cn/fase/EN/Y2019/V6/I1/61
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Chengcheng ZHAO
Junsong SHI
Rong ZHOU
Ranbiao MAI
Lvhua LUO
Xiaoyan HE
Hongmei JI
Gengyuan CAI
Dewu LIU
Enqin ZHENG
Zhenfang WU
Zicong LI
Enucleation method Number of manipulated/successfully enucleated oocytes Enucleation rate/%
BA-B 384/319 83.07**
SIS-A 322/306 95.03**
Tab.1  Comparison of the success rates of BA-B and SIS-A enucleation
Enucleation method SCNT embryos Cleaved/% Blastocysts/% Total number of cells per blastocyst
BA-B 240 183 (76.25) 44 (18.33) 40±3.34
SIS-A 279 208 (74.55) 37 (13.26) 47±4.10
Tab.2  Comparison of in vitro development rate of pig SCNT embryos generated by BA-B and SIS-A enucleation
Enucleation method Total number of transferred SCNT embryos Total number/pregnant/farrowed recipients Pregnancy/farrowing rate/% Total number of cloned piglets born Development rate/%
BA-B 7469 33/23/14 69.70/42.42 64 0.86
SIS-A 6779 33/23/14 69.70/42.42 75 1.11*
Tab.3  Comparison of in vivo development rate of pig SCNT embryos generated by the BA-B and SIS-A enucleation
Enucleation method SCNT embryos Cleaved/% Blastocysts/% Total number of cells per blastocyst
BA-B 429 326 (75.99) 94 (21.91) 36±1.77*
BAPPS-B 501 362 (72.26) 89 (17.76) 41±1.69*
Tab.4  Comparison of in vitro development rate of pig SCNT embryos generated by the BA-B and BAPPS-B enucleation
Enucleation method Number of manipulated /successfully enucleated oocytes Total number of transferred SCNT embryos Total number/pregnant/farrowed recipients Pregnancy/farrowing rate/% Total number of born cloned piglets Development rate/%
BA-B 8023/not determined 7038 31/18/7 58.06/22.58* 24 0.34**
BAPPS-B 9049/7234 6735 31/21/15 67.74/48.39* 55 0.82**
Tab.5  Comparison of in vivo development rate of pig SCNT embryos generated by the BA-B an BAPPS-B enucleation
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